Alkane diol derivatives as therapeutic agents for the treatment of bone conditions

ABSTRACT

The present invention pertains to certain alkane diol derivatives (including, e.g., mono- and di-esters) of the formula R 1 —O-A-O—R 2 , wherein: A is a C 2-10  alkylene group; R 1  is independently a first hydroxy protecting group (e.g., an ester group); and, R 2  is independently —H or a second hydroxy protecting group (e.g., an ester group); and pharmaceutically acceptable salts, solvates, amides, esters, ethers, chemically protected forms, and prodrug thereof, which, inter alia, inhibit osteoclast survival, formation, and/or activity; and/or inhibit bone resorption. The present invention also pertains to pharmaceutical compositions comprising such compounds, and the use of such compounds and compositions, both in vitro and in vivo, to inhibit osteoclast survival, formation, and/or activity, and to inhibit conditions mediated by osteoclasts and/or characterised by bone resorption, such as osteoporosis, rheumatoid arthritis, cancer associated bone disease, Paget&#39;s disease, and the like; and/or conditions associated with inflammation or activation of the immune system.

This application is the US national phase of international applicationPCT/GB02/04933 filed in English on 31 Oct. 2002, which designated theUS. PCT/GB02/04933 claims priority to GB Application No. 0126157.7 filed31 Oct. 2001. The entire contents of these applications are incorporatedherein by reference.

RELATED APPLICATION

This application is related to (and where permitted by law, claimspriority to) United Kingdom patent application number GB 0126157.7 filed31 Oct. 2001, the contents of which are incorporated herein by referencein their entirety.

TECHNICAL FIELD

The present invention pertains generally to the field of therapeuticcompounds for treating bone conditions, and more specifically to certainalkane diol derivatives which, inter alia, inhibit osteoclast survival,formation, and/or activity; and/or inhibit bone resorption. The presentinvention also pertains to pharmaceutical compositions comprising suchcompounds, and the use of such compounds and compositions, both in vitroand in vivo, to inhibit osteoclast survival, formation, and/or activity,and to inhibit conditions mediated by osteoclasts and/or characterisedby bone resorption, such as osteoporosis, rheumatoid arthritis, cancerassociated bone disease, Paget's disease, and the like; and/orconditions associated with inflammation or activation of the immunesystem.

BACKGROUND

Throughout this specification, including the claims which follow, unlessthe context requires otherwise, the word “comprise,” and variations suchas “comprises” and “comprising,” will be understood to imply theinclusion of a stated integer or step or group of integers or steps butnot the exclusion of any other integer or step or group of integers orsteps.

It must be noted that, as used in the specification and the appendedclaims, the singular forms “a,” “an,” and “the” include plural referentsunless the context clearly dictates otherwise. Thus, for example,reference to “a pharmaceutical carrier” includes mixtures of two or moresuch carriers, and the like.

Ranges are often expressed herein as from “about” one particular value,and/or to “about” another particular value. When such a range isexpressed, another embodiment includes from the one particular valueand/or to the other particular value. Similarly, when values areexpressed as approximations, by the use of the antecedent “about,” itwill be understood that the particular value forms another embodiments.

Functions of Bone

The function of bone is to provide mechanical support for joints,tendons and ligaments, to protect vital organs from damage and to act asa reservoir for calcium and phosphate in the preservation of normalmineral homeostasis. Diseases of bone compromise these functions,leading to clinical problems such as bone pain, bone deformity, fractureand abnormalities of calcium and phosphate homeostasis.

Types of Bone

The normal skeleton contains two types of bone; cortical or compactbone, which makes up most of shafts (diaphysis) of the long bones suchas the femur and tibia, and trabecular or spongy bone which makes upmost of the vertebral bodies and the ends of the long bones.

Trabecular bone has a greater surface area than cortical bone andbecause of this is remodeled more rapidly. This means that conditionsassociated with increased bone turnover tend to affect trabecular bonemore quickly and more profoundly than cortical bone. Cortical bone isarranged in so-called Haversian systems which consists of a series ofconcentric lamellae of collagen fibres surrounding a central canal thatcontains blood vessels. Nutrients reach the central parts of the bone byan interconnecting system of canaliculi that run between osteocytesburied deep within bone matrix and lining cells on the bone surface.Trabecular bone has a similar structure, but here the lamellae run inparallel to the bone surface, rather than concentrically as in corticalbone.

Bone Composition

The organic component of bone matrix comprises mainly of type Icollagen; a fibrillar protein formed from three protein chains, woundtogether in a triple helix. Collagen type I is laid down by bone formingcells (osteoblasts) in organised parallel sheets (lamellae) andsubsequently the collagen chains become cross-linked by specialisedcovalent bonds which help to give bone its tensile strength. When boneis formed rapidly (for example in Paget's disease, or in bonemetastases), the lamellae are laid down in a disorderly fashion givingrise to “woven bone”, which is mechanically weak and easily fractured.Bone matrix also contains small amounts of other collagens and severalnon-collagenous proteins and glycoproteins. Some of these, such asosteocalcin, are specific to bone, whereas others, such as osteopontinand fibronectin and various peptide growth factors are also found inother connective tissues. The function of non-collagenous bone proteinsis unclear, but it is thought that they are involved in mediating theattachment of bone cells to bone matrix, and in regulating bone cellactivity during the process of bone remodelling. The organic componentof bone forms a framework upon which mineralisation occurs. During boneformation, osteoblasts lay down uncalcified bone matrix (osteoid) whichcontains the components described above and small amounts of otherproteins, which are adsorbed from extracellular fluid. After a lag phaseof about 10 days, the matrix becomes mineralised, as hydroxyapatite((Ca₁₀ (PO₄)₆ (OH)₂) crystals are deposited in the spaces betweencollagen fibrils. Mineralisation confers upon bone the property ofmechanical rigidity, which complements the tensile strength, andelasticity derived from bone collagen.

Bone Cell Function and Bone Remodelling

The mechanical integrity of the skeleton is maintained by the process ofbone remodelling, which occurs throughout life, in order that damagedbone can be replaced by new bone. Remodelling can be divided into fourphases; resorption; reversal, formation and quiescence (see, e.g.,Raisz, 1988; Mundy, 1996). At any one time approximately 10% of bonesurface in the adult skeleton is undergoing active remodeled whereas theremaining 90% is quiescent.

Osteoclast Formation and Differentiation

Remodelling commences with attraction of bone resorbing cells(osteoclasts) to the site, which is to be resorbed. These aremultinucleated phagocytic cells, rich in the enzyme tartrate-resistantacid phosphatase, which are formed by fusion of precursors derived fromthe cells of monocyte/macrophage lineage. Recent work has identifiedseveral molecules that are of key importance in the regulation ofosteoclast differentiation (see, e.g., Ralston, 1997). The transcriptionfactor PU-1 which is expressed in early osteoclast precursors isnecessary for the initial stages of osteoclast and monocytedifferentiation, whereas other transcription factors including c-fos andNFkB play an essential role in stimulating differentiation of committedprecursors to mature osteoclasts. Osteoclast formation and activation isalso dependent on close contact between osteoclast precursors and bonemarrow stromal cells. Stromal cells secrete the cytokine M-CSF(macrophage colony stimulating factor), which is essential fordifferentiation of both osteoclasts and macrophages from a commonprecursor. Stromal cells also express a molecule called RANK ligand(RANKL) on the cell surface, which interacts with another cell surfacereceptor present on osteoclast precursors called RANK (BeceptorActivator of Nuclear Factor Kappa B) to promote differentiation ofosteoclast precursors to mature osteoclasts. The RANK-RANKL interactionis blocked by another molecule called Osteoprotegerin (OPG), which is a“decoy” ligand for RANK and which acts a potent inhibitor of osteoclastformation (see, e.g., Kong et al., 1999; Yasuda et al., 1998). Recentwork suggests that many of the factors that promote osteoclast formationand bone resorption do so by regulating expression of these molecules.

Mature osteoclasts form a tight seal over the bone surface and resorbbone by secreting hydrochloric acid and proteolytic enzymes through the“ruffled border” into a space beneath the osteoclast (Howship's lacuna).Formation of this ruffled border is critically dependent on the presenceof c-src, a cell membrane associated signalling protein. Thehydrochloric acid secreted by osteoclasts dissolves hydroxyapatite andallows proteolytic enzymes (mainly Cathepsin K and matrixmetalloproteinases) to degrade collagen and other matrix proteins.Molecules which have been identified as being important in regulatingosteoclast activity include; carbonic anhydrase II (Ca-II) whichcatalyses formation of hydrogen ions within osteoclasts; TCIRG1, whichencodes a subunit of the osteoclast proton pump, and Cathepsin K whichdegrades collagen and other non-collagenous proteins. Deficiency ofthese proteins causes osteopetrosis which is a disease associated withincreased bone density and osteoclast dysfunction. After resorption iscompleted osteoclasts undergo programmed cell death (apoptosis), in theso-called reversal phase which heralds the start of bone formation. Ithas recently been discovered that many of the drugs, which are usedclinically to inhibit bone resorption, such as bisphosphonates andoestrogen do so by promoting osteoclast apoptosis (see, e.g., Hughes etal., 1997).

Osteoblast Formation and Differentiation

Bone formation begins with attraction of osteoblast precursors, whichare derived from mesenchymal stem cells in the bone marrow, to the bonesurface. Although these cells have the potential to differentiate intomany cell types including adipocytes, myocytes, and chondrocytes it isnow known that the key trigger for osteoblast differentiation isexpression of a regulatory molecule called Cbfa1 in pre-osteoblasts(see, e.g., Rodan et al., 1997). Cbfa1 is a transcription factor thatactivates co-ordinated expression of genes characteristic of theosteoblast phenotype such as osteocalcin, type I collagen and alkalinephosphatase. In contrast, expression of the transcription factor PPARgpromotes the cells towards adipocyte differentiation. It is currentlythought that some cases of osteoporosis may occur because there is animbalance between the rate of osteoblast and adipocyte differentiationin bone. Mature osteoblasts are plump cuboidal cells, which areresponsible for the production of bone matrix. They are rich in theenzyme alkaline phosphatase and the protein osteocalcin, which are usedclinically as serum markers of osteoblast activity. Osteoblasts lay downbone matrix which is initially unmineralised (osteoid), but whichsubsequently becomes calcified after about 10 days to form mature bone.During bone formation, some osteoblasts become trapped within the matrixand differentiate into osteocytes, whereas others differentiate intoflattened “lining cells” which cover the bone surface. Osteocytesconnect with one another and with lining cells on the bone surface by anintricate network of cytoplasmic processes, running through cannaliculiin bone matrix. Osteocytes appear to act as sensors of mechanical strainin the skeleton, and release signalling molecules such as prostaglandinsand nitric oxide (NO), which modulate the function of neighbouring bonecells.

Regulation of Bone Remodelling

Bone remodelling is a highly organised process, but the mechanisms whichdetermine where and when remodelling occurs are poorly understood.Mechanical stimuli and areas of micro-damage are likely to be importantin determining the sites at which remodelling occurs in the normalskeleton. Increased bone remodelling may result from local or systemicrelease of inflammatory cytokines like interleukin-1 and tumour necrosisfactor in inflammatory diseases. Calciotropic hormones such asparathyroid hormone (PTH) and 1,25-dihydroxyvitamin D, act together toincrease bone remodelling on a systemic basis allowing skeletal calciumto be mobilised for maintenance of plasma calcium homeostasis. Boneremodelling is also increased by other hormones such as thyroid hormoneand growth hormone, but suppressed by oestrogen, androgens andcalcitonin.

Common Bone Diseases

Osteoporosis is a common disease characterized by reduced bone density,deterioration of bone tissue and increase risk of fracture. Many factorscontribute to the pathogenesis of osteoporosis including poor diet, lackof exercise, smoking and excessive alcohol intake. Osteoporosis may alsoarise in association with inflammatory diseases such as rheumatoidarthritis, endocrine diseases such as thyrotoxicosis and with certaindrug treatments such as glucocorticoids. However one of the mostimportant factors in the pathogenesis of osteoporosis is heredity.

Paget's disease of bone is a common condition of unknown cause,characterized by increased bone turnover and disorganized boneremodeling, with areas of increased osteoclastic and osteoblastactivity. Although Pagetic bone is often denser than normal, theabnormal architecture causes the bone to be mechanically weak, resultingin bone deformity and increased susceptibility to pathological fracture.

Multiple Myeloma is a cancer of plasma cells. In contrast to most otherhaematological malignancies, the tumour cells do not circulate in theblood, but accumulate in the bone marrow where they give rise to highlevels of cytokines that activate osteoclastic bone resorption (e.g.,interleukin-6). The disease accounts for approximately 20% of allhaematological cancers and is mainly a disease of elderly people.

Bone Resorption Inhibitors

Several common diseases, such as osteoporosis and rheumatoid arthritis,are characterised by bone loss due to excess bone resorption byosteoclasts. At present the most commonly used types of drugs used tosuppress osteoclast activity in these diseases are bisphophonates (BPs)and non-steroidal anti-inflammatory drugs (NSAIDs).

Bisphosphonates (also know as diphosphonates) are an important class ofdrugs used in the treatment of bone diseases involving excessive bonedestruction or resorption, e.g., Paget's disease, tumour-associatedosteolysis, and post-menopausal osteoporosis. Bisphosphonates arestructural analogues of naturally occurring pyrophosphate. Whereaspyrophosphate consists of two phosphate groups linked by an oxygen atom(P—O—P), bisphosphonates have two phosphate groups linked by a carbonatom (P—C—P). This makes bisphosphonates very stable and resistant todegradation. Furthermore, like pyrophosphate, bisphosphonates have veryhigh affinity for calcium and therefore target to bone mineral in vivo.The carbon atom that links the two phosphate groups has two side chainsattached to it, which can be altered in structure. This gives rise to amultitude of bisphosphonate compounds with different anti-resorptivepotencies. Bone resorption is mediated by highly specialised,multinucleated osteoclast cells. Bisphosphonate drugs specificallyinhibit the activity and survival of these cells. Firstly, afterintravenous or oral administration, the bisphosphonates are rapidlycleared from the circulation and bind to bone mineral. As the mineral isthen resorbed and dissolved by osteoclasts, it is thought that the drugis released from the bone mineral and is internalised by osteoclasts.Intracellular accumulation of the drugs inhibits the ability of thecells to resorb bone (probably by interfering with signal transductionpathways or cellular metabolism) and causes osteoclast apoptosis.

NSAIDs are widely used in the treatment of inflammatory diseases, butoften cause severe gastro-intestinal (GI) side effects. NSAIDs developedby Nicox SA (Sophia Antipolis, France), that contain a nitric oxide(NO)-donor group (NO-NSAID) exhibit anti-inflammatory properties withoutcausing GI side effects. An example of such a compound is HCT 1026,which is a nitrosylated derivative of the NSAID flurbiprofen (see, forexample, Armour et al., 2001).

There is a recognized need for more and better treatments for these andother bone-related diseases, which offer, for example, one or more thefollowing benefits:

-   (a) improved activity;-   (b) improved efficacy;-   (c) improved specificity;-   (d) reduced toxicity (e.g., cytotoxicity);-   (e) complement the activity of other treatments (e.g.,    chemotherapeutic agents);-   (f) reduced intensity of undesired side-effects;-   (g) fewer undesired side-effects;-   (h) simpler methods of administration (e.g., route, timing,    compliance);-   (i) reduction in required dosage amounts;-   (j) reduction in required frequency of administration;-   (k) increased ease of synthesis, purification, handling, storage,    etc.;-   (l) reduced cost of synthesis, purification, handling, storage, etc.

Thus, one aim of the present invention is the provision of activecompounds which offer one or more of the above benefits.

SUMMARY OF THE INVENTION

One aspect of the invention pertains to a method of inhibitingosteoclast survival, formation, and activity, in vitro or in vivo,comprising contacting an osteoclast with an effective amount of anactive compound, as described herein.

Another aspect of the invention pertains to a method of inhibiting boneresorption, in vitro or in vivo, comprising contacting cells in the bonemicroenvironment with a therapeutically-effective amount of an activecompound, as described herein.

Another aspect of the present invention pertains to a method for thetreatment of a condition mediated by osteoclasts and/or characterised bybone resorption, as described herein, comprising administering to asubject suffering from said condition a therapeutically-effective amountof an active compound, as described herein, preferably in the form of apharmaceutical composition.

Another aspect of the present invention pertains to a method for thetreatment of a condition associated with inflammation or activation ofthe immune system, as described herein, comprising administering to asubject suffering from said condition a therapeutically-effective amountof an active compound, as described herein, preferably in the form of apharmaceutical composition.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition mediated by osteoclasts and/orcharacterised by bone resorption, as described herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition mediated by osteoclasts, asdescribed herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition characterised by bone resorption, asdescribed herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of osteoporosis, rheumatoid arthritis, cancerassociated bone disease, or Paget's disease.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition associated with inflammation oractivation of the immune system, as described herein.

Another aspect of the present invention pertains to an active compoundas described herein for use in a method of treatment of the human oranimal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionmediated by osteoclasts and/or characterised by bone resorption, asdescribed herein, of the human or animal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionmediated by osteoclasts, as described herein, of the human or animalbody by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditioncharacterised by bone resorption, as described herein, of the human oranimal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment of osteoporosis,rheumatoid arthritis, cancer associated bone disease, or Paget's diseaseof the human or animal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionassociated with inflammation or activation of the immune system, asdescribed herein, of the human or animal body by therapy.

Another aspect of the invention pertains to active compounds,specifically, certain alkane diol derivatives (e.g., esters of alkanediols) as described herein.

Another aspect of the invention pertains to a composition comprising anactive compound as described herein and a pharmaceutically acceptablecarrier or diluent.

Another aspect of the present invention pertains to a kit comprising (a)an active compound, as described herein, preferably provided as apharmaceutical composition and in a suitable container and/or withsuitable packaging; and (b) instructions for use, for example, writteninstructions on how to administer the active compound.

Another aspect of the present invention pertains to compounds obtainableby a method of synthesis as described herein, or a method comprising amethod of synthesis as described herein.

Another aspect of the present invention pertains to compounds obtainedby a method of synthesis as described herein, or a method comprising amethod of synthesis as described herein.

Another aspect of the present invention pertains to novel intermediates,as described herein, which are suitable for use in the methods ofsynthesis described herein.

Another aspect of the present invention pertains to the use of suchnovel intermediates, as described herein, in the methods of synthesisdescribed herein.

As will be appreciated by one of skill in the art, features andpreferred embodiments of one aspect of the invention will also pertainto other aspect of the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph of macrophage viability, as measured by the MTT andAlamar Blue macrophage J774 viability assays, expressed as % of control,after 72 hour exposure to ABD-0028 (4T) (“28”) and ABD-0042 (4BPA)(“42”) as a function of concentration of compound.

FIG. 2 is a bar graph showing the number of osteoclasts, expressed as a% of control value, after three days exposure to compound, for themurine co-culture system, for several examples of biphenylcarboxy (BP),trityl (T), and ibuprofenyl (1) compounds: ABD-0057 (3BP), ABD-0056(4BP), ABD-0055 (5BP), ABD-0054 (6BP), ABD-0028 (4T), ABD-0037 (31),ABD-0036 (41), ABD-0038 (51), and ABD-0039 (61). All compounds weretested at 100 μM concentration. Each value represents an average of 3experiments, each of which had 5 data points.

FIG. 3 is a graph showing the effects of compounds on osteoclast andJ774 survival, for both the murine co-culture system (A) and the MTTmacrophage J774 viability assay (B), for several examples ofbiphenylcarboxy (BP), trityl (T), and ibuprofenyl (I) compounds, andsome others. For co-culture (A), the graph shows number of osteoclasts,expressed as a % of control value. For macrophages (B), the graph showsviability as measured by the MTT assay, expressed as % of control. Allcompounds were tested at 100 μM concentration.

FIG. 4 is a graph of macrophage viability, as measured by the MTTmacrophage J774 viability assay, expressed as % of control, after 24hour exposure to compound, for several examples of biphenylcarboxy (BP)compounds: ABD-0053 (BuBP), ABD-0057 (38P), ABD-0056 (4BP), ABD-0055(5BP), ABD-0054 (6BP). Unless shown, error bars are less than 20%.

FIG. 5 is a graph of macrophage viability, as measured by the MTTmacrophage J774 viability assay, expressed as % of control, after 24hour exposure to compound, for several examples of ibuprofenyl (I)compounds: ABD-0035 (Bul), ABD-0037 (31), ABD-0036 (41), ABD-0038 (51),ABD-0039 (61). Unless shown, error bars are less than 20%.

FIG. 6 is a graph of macrophage viability, as measured by the MTTmacrophage J774 viability assay, expressed as % of control, after 24hour exposure to compound, for several examples of trityl (T) compounds:ABD-0028 (4T), ABD-0030 (5T), ABD-0031 (6T). Unless shown, error barsare less than 20%.

FIG. 7 is a graph of macrophage viability, as measured by the MTTmacrophage J774 viability assay, expressed as % of control, after 24hour exposure to compound, for several examples of biphenylacetyl (BPA)compounds: ABD-0040 (BuBPA), ABD-0041 (3BPA), ABD-0042 (4BPA), ABD-0043(5BPA), ABD-0044 (6BPA). Unless shown, error bars are less than 20%.

FIG. 8 is a graph of macrophage viability, as measured by the MTTmacrophage J774 viability assay, expressed as % of control, after 24hour exposure to compound, for several examples of butanediol compounds:ABD-0042 (4BPA), ABD-0028 (4T), ABD-0056 (4BP), ABD-0036 (41).

FIG. 9 is a graph of macrophage viability, as measured by the AlamarBlue macrophage J774 viability assay, expressed as % of control, after72 hour exposure to compound, for several examples of butanediolcompounds: ABD-0042 (4BPA), ABD-0028 (4T), ABD-0056 (4BP), ABD-0036(41).

FIG. 10 is a graph of macrophage viability, as measured by the AlamarBlue macrophage J774 viability assay, expressed as % of control, after72 hour exposure to compound, for ABD-0098 (“2NO₂”), ABD-0100 (“4F”),ABD-0099 (“2F”), ABD-0089 (“Xyl”), and ABD-0102 (“4Br”).

FIG. 11 is a graph of macrophage viability, as measured by the AlamarBlue macrophage J774 viability assay, expressed as % of control, after72 hour exposure to compound, for ABD-0072 (“OH”), ABD-0089(“Dimethyl”), ABD-0070 (“Methyl”), ABD-0094 (“Ethyl”), and ABD-0097(“Methoxy”).

FIG. 12 is a graph of macrophage viability, as measured by the AlamarBlue macrophage J774 viability assay, expressed as % of control, after72 hour exposure to compound, for ABD-0085 (10F) and ABD-0077 (5F).

FIG. 13 is a graph showing the effects of compounds in the rabbitosteoclast culture system, and is a plot of the number of rabbitosteoclasts, expressed as a % of control value, after three daysexposure to compound, as a function of compound concentration, forseveral examples of biphenylcarboxy (BP) derivatives: ABD-0053 (BuBP),ABD-0057 (3BP), ABD-0056 (4BP), ABD-0055 (5BP), ABD-0054 (6BP).

FIG. 14 is a graph showing the effects of compounds in the rabbitosteoclast culture system, and is a plot of resorption pit areaexpressed as a % of control, after three days exposure to compound, as afunction of compound concentration, for several examples ofbiphenylcarboxy (BP) derivatives: ABD-0053 (BuBP), ABD-0057 (3BP),ABD-0056 (4BP), ABD-0055 (5BP), ABD-0054 (6BP). Each value represents anaverage of 3 experiments, each of which had 5 data points.

FIG. 15 is a graph showing the effects of compounds in the murineco-culture system, and is a plot of the number of murine osteoclasts,expressed as a % of control value as a function of compoundconcentration, for ABD-0056 (4BP). The compound is added at Day 2 andthere is a complete medium refresh after Day 4 to remove the compound.The experiment is terminated at Day 10 and the number of osteoclastsascertained by TRAcP staining. Each value represents an average of 3experiments, each of which had 5 data points.

FIG. 16 is a graph showing the effects of compounds in the murineco-culture system, and is a plot of resorption pit area expressed as a %of control value as a function of compound concentration, for ABD-0056(4BP). The compound is added at Day 2 and there is a complete mediumrefresh after Day 4 to remove the compound. The experiment is terminatedat Day 10 and the amount of resorption measured by reflected lightmicroscopy. Each value represents an average of 3 experiments, each ofwhich had 5 data points.

FIG. 17 is a graph showing the effects of compounds in the murineco-culture system, and is a plot of the number of murine osteoclasts,expressed as a % of control value, after three days exposure to compoundadded at Day 7, as a function of compound concentration, for severalexamples of biphenylcarboxy (BP) derivatives: ABD-0053 (BuBP), ABD-0057(3BP), ABD-0056 (4BP), ABD-0055 (5BP), ABD-0054 (6BP).

FIG. 18 is a graph showing the effects of compounds in the murineco-culture system, and is a plot of resorption pit area expressed as a %of control, after three days exposure to compound added at Day 7, as afunction of compound concentration, for several examples ofbiphenylcarboxy (BP) derivatives: ABD-0053 (BuBP), ABD-0057 (3BP),ABD-0056 (4BP), ABD-0055 (5BP), ABD-0054 (6BP). Each value represents anaverage of 3 experiments, each of which had 5 data points.

FIG. 19 is a graph of osteoclast number and resorption pit area for themurine co-culture system, where ABD-0056 (4BP) was added at Day 7, andincubation continued until Day 10 (mature osteoclasts).

DETAILED DESCRIPTION OF THE INVENTION

One aspect of the present invention pertains to compounds which may bedescribed as alkane diol derivatives, and their surprising andunexpected osteoclast-inhibitory and resorption-inhibitory effects.

Alkane Diol Derivatives

One aspect of the present invention pertains to compounds which may bedescribed as derivatives of the alkane diols described above (i.e.,alkane diol derivatives), and which have the following formula:

wherein:

-   -   A is a C₂₋₁₀alkylene group;    -   R¹ is independently a first hydroxy protecting group; and,    -   R² is independently —H or a second hydroxy protecting group;    -   and pharmaceutically acceptable salts, solvates, amides, esters,        ethers, chemically protected forms, or prodrugs thereof.

In one embodiment, R² is not —H, and the compound is di-protected.

In one embodiment, R² is not —H, and R¹ and R² are the same.

In one embodiment, R² is not —H, and R¹ and R² are different.

In one embodiment, R² is —H, the compound is mono-protected, and has thefollowing formula:

Alkane Diols

The compounds of the present invention may be described as alkane diolderivatives. In this context, the alkane diols have the followingformula:HO-A-OH  (3)wherein A is a C₂₋₁₀alkylene group, and is optionally substituted.The Alkylene Group, A

The alkylene group, A, is a C₂₋₁₀alkylene group, and is optionallysubstituted.

The term “C₂₋₁₀alkylene,” as used herein, pertains to a bidentate moietyobtained by removing two hydrogen atoms, either both from the samecarbon atom, or one from each of two different carbon atoms, of ahydrocarbon compound having from 2 to 10 carbon atoms, which may bealiphatic or alicyclic, or a combination thereof, and which may besaturated, partially unsaturated, or fully unsaturated. The prefix(i.e., “C₂₋₁₀”) denotes the number of carbon atoms in the moiety.

In one embodiment, the alkylene group is unsubstituted.

In one embodiment, the two hydrogen atoms are removed from differentcarbon atoms.

In one embodiment, the two hydrogen atoms are removed from differentcarbon atoms, and these carbon atoms are not adjacent.

In one embodiment, the alkylene group is C₃₋₁₀alkylene group.

In one embodiment, the alkylene group is C₄₋₁₀alkylene group.

In one embodiment, the alkylene group is C₂₋₈alkylene group.

In one embodiment, the alkylene group is C₃₋₈alkylene group.

In one embodiment, the alkylene group is C₄₋₈alkylene group.

In one embodiment, the alkylene group is C₂₋₇alkylene group.

In one embodiment, the alkylene group is C₃₋₇alkylene group.

In one embodiment, the alkylene group is C₄₋₇alkylene group.

In one embodiment, the alkylene group is C₂₋₆alkylene group.

In one embodiment, the alkylene group is C₃₋₆alkylene group.

In one embodiment, the alkylene group is C₄₋₆alkylene group.

In one embodiment, the alkylene group is C₃alkylene group.

In one embodiment, the alkylene group is C₄alkylene group.

In one embodiment, the alkylene group is C₅alkylene group.

In one embodiment, the alkylene group is C₆alkylene group.

In one embodiment, the alkylene group is an aliphatic group.

In one embodiment, the alkylene group is a branched group.

In one embodiment, the alkylene group is a linear group.

In one embodiment, the alkylene group is a partially unsaturatedaliphatic group.

In one embodiment, the alkylene group is a fully saturated aliphaticgroup.

In one embodiment, the alkylene group is a partially unsaturatedbranched group. Examples of such groups include, but are not limited to,the following:

-   —C(Me)=CH—, —CH═C(Me)—, —C(Me)=C(Me)—,-   —C(Et)=CH—, —CH═C(Et)-, —C(Et)=C(Et)-,-   —C(Me)=CH—CH₂—, —CH═C(Me)-CH₂—, —CH═CH—CH(Me)—,-   —C(Et)=CH—CH₂—, —CH═C(Et)-CH₂—, —CH═CH—CH(Et)-,-   —C(Me)=CH—CH₂CH₂—, —CH═C(Me)-CH₂CH₂—, —CH═CH—CH(Me)CH₂—,-   —C(Et)=CH—CH₂CH₂—, —CH═C(Et)-CH₂CH₂—, and —CH═CH—CH(Et)CH₂—.

In one embodiment, the alkylene group is a fully saturated branchedgroup.

Examples of such groups include, but are not limited to, the following:

-   —CH(Me)—, —CH(Et)-,-   —CH(Me)CH₂—, —CH(Et)CH₂—, —CH₂CH(Me)—, —CH₂CH(Et)-,-   —CH(Me)CH₂CH₂—, —CH₂CH(Me)CH₂—, —CH₂CH₂CH(Me)—,-   —CH(Et)CH₂CH₂—, —CH₂CH(Et)CH₂—, —CH₂CH₂CH(Et)-,-   —CH(Me)CH₂CH₂CH₂—, —CH₂CH(Me)CH₂CH₂—,-   —CH(Et)CH₂CH₂CH₂—, and —CH₂CH(Et)CH₂CH₂—.

In one embodiment, the alkylene group is a partially unsaturated lineargroup.

Examples of such groups include, but are not limited to, the following:

-   —CH═CH— (vinylene),-   —CH═CH—CH₂—, —CH₂—CH═CH—,-   —CH═CH—CH₂—CH₂—, —CH₂—CH—CH═CH—, —CH═CH—CH═CH—,-   —CH═CH—CH₂—CH₂—CH₂—, —CH═CH—CH═CH—CH₂—, —CH═CH—CH₂—CH═CH—,-   —CH═CH—CH═CH—CH₂—CH₂—, and —CH═CH—CH₂—CH₂—CH═CH—.

In one embodiment, the alkylene group is a fully saturated linear group.

Examples of such groups include, but are not limited to, groups of theformula —(CH₂)_(n)— where n is an integer from 2 to 10, for example,—(CH₂)₂— (ethylene), —(CH₂)₃— (propylene), —(C H₂)₄— (butylene),—(CH₂)₅— (pentylene), —(CH₂)₆-(hexylene), —(CH₂)₇— (heptylene), —(CH₂)₈—(octylene), —(CH₂)₉— (nonylene), and —(CH₂)₁₀— (decylene).

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 3 to 10.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 4 to 10.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 2 to 8.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 3 to 8.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 4 to 8.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 2 to 7.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 3 to 7.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 4 to 7.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 2 to 6.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 3 to 6.

In one embodiment, the alkylene group is —(CH₂)_(n)— where n is aninteger from 4 to 6.

In one embodiment, the alkylene group is —(CH₂)₃—.

In one embodiment, the alkylene group is —(CH₂)₄—.

In one embodiment, the alkylene group is —(CH₂)₅—.

In one embodiment, the alkylene group is —(CH₂)₆—.

In one especially preferred embodiment, the alkylene group is —(CH₂)₄—:

In one embodiment, the alkylene group is substituted, for example, withone or more substituents selected from: halogen, hydroxy, ether (e.g.,C₁₋₇alkoxy), amino, and amido.

In one embodiment, the alkylene group is substituted, for example, withone or more substituents selected from: —F, —Cl, —Br, and —I.

In one embodiment, the alkylene group is substituted, for example, withone or more —F groups.

In one embodiment, the alkylene group is:

The Hydroxy Groups, —OH, of the Alkane Diol

The hydroxy groups, —OH, of the alkane diol may be primary, secondary,or tertiary.

In one embodiment, the hydroxy groups are primary or secondary.

In one embodiment, at least one of the hydroxy groups is a primaryhydroxy group.

In one embodiment, each of the hydroxy groups is a primary hydroxygroup.

In one embodiment, the hydroxy groups are not geminal.

In one embodiment, the hydroxy groups are not geminal, and are notvicinal.

Some Preferred Alkane Diols

In one embodiment, the alkane diol has the formula HO—(CH₂)_(n)—OH,where n is an integer from 2 to 10.

HO—(CH₂)₂—OH

1,2-ethanediol HO—(CH₂)₃—OH

1,3-propanediol HO—(CH₂)₄—OH

1,4-butanediol HO—(CH₂)₅—OH

1,5-pentanediol HO—(CH₂)₆—OH

1,6-hexanediol HO—(CH₂)₇—OH

1,7-heptanediol HO—(CH₂)₈—OH

1,8-octanediol HO—(CH₂)₉—OH

1,9-nonanediol HO—(CH₂)₁₀—OH

1,10-decanediol

In one embodiment, the alkane diol is 1,3-propanediol, 1,4-butanediol,1,5-pentanediol, or 1,6-hexanediol.

In one embodiment, the alkane diol is 1,4-butanediol, 1,5-pentanediol,or 1,6-hexanediol.

In one embodiment, the alkane diol is 1,4-butanediol or 1,6-hexanediol.

In one especially preferred embodiment, the alkane diol is1,4-butanediol:

Hydroxy Protecting Groups

Without wishing to be bound by any particular theory, it is believedthat the hydroxy protecting group (or, if there are two such groups, oneof them, or each of them) additionally conveys one or more advantageousproperties (e.g., solubility, lipophilicity, targeting) to the resultingcompound.

For example, the compound is preferably sufficiently soluble in therelevant media (e.g., water, culture media, in vivo) so that, forexample, its beneficial therapeutic properties may be realised. Thus, inone embodiment, the hydroxy protecting group (or, if there are two suchgroups, one of them, or each of them) is selected so that the resultingcompound has acceptable solubility properties.

Similarly, the compound is preferably readily able to cross cellmembranes, and more specifically, the membranes of cells for whichtreatment is sought. Thus, in one embodiment, the hydroxy protectinggroup (or, if there are two such groups, one of them, or each of them)is selected to be, or to comprise, a hydrophobic group, so that theresulting compound is better able to able to cross cell membranes.

Also, in one embodiment, the compound is targeted to bone and/or thebone environment. Thus, in one embodiment, the hydroxy protecting group(or, if there are two such groups, one of them, or each of them) isselected to be, or to comprise, a bone-targeting group.

The term “bone-targeting group,” as used herein, pertains to a chemicalmoiety which has an affinity for bone and/or the bone environment, andwhich, when attached to compound, acts as a targeting moiety, and soaids in the delivery of that compound to the bone and/or boneenvironment. Examples of such bone-targeting groups include phosphonicacid groups, and salts, esters, and amides thereof, as discussed below.

Esters as Hydroxy Protecting Groups

In one embodiment, the hydroxy protecting group (or, if there are twosuch groups, one of them, or each of them) is an acyl group (i.e.,R^(A)—C(═O)—), and the protected hydroxy group is an ester group (i.e.,R^(A)—C(═O)—). More specifically, the hydroxy group is protected as anester group.

In one embodiment, R² is not —H, and each of the hydroxy protectinggroups are acyl groups (R^(A1)—C(═O)— and R^(A2)—C(═O)—, respectively),which may be the same or different, and the compound has the followingformula (also referred to herein as a “di-ester” of the alkane diol):

In one embodiment, R² is not —H, and R^(A1) and R^(A2) are the same.

In one embodiment, R² is not —H, and R^(A1) and R^(A2) are different.

In one embodiment, R² is —H, the hydroxy protecting group is an acylgroup (i.e., R^(A1)—C(═O)—), and the compound has the following formula(also referred to herein as a “mono-ester” of the alkane diol):

Acyl Groups, R^(A1)—C(═O)— and R^(A2)—C(═O)—

In one embodiment, R^(A1) is independently a C₁₋₇alkyl group, aC₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, and is optionallysubstituted; and, R^(A2), if present, is independently, a C₁₋₇alkylgroup, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, and isoptionally substituted.

In one especially preferred embodiment, R^(A1) is independently aC₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, and is optionallysubstituted

In one especially preferred embodiment, R^(A1) is independently anoptionally substituted C₅₋₂₀aryl group, as described below.

As discussed above, one or both of the moieties R^(A1) and R^(A2) may beselected so that the resulting compound (a) has improved solubility, (b)is better able to cross cell membranes, (c) is, or comprises, abone-targeting moiety; or a combination thereof

R^(A1) as Optionally Substituted Aryl

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted C₅₋₂₀aryl group.

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted phenyl, naphthyl, pyridyl, furanyl,thiofuranyl, indolyl, pyrrolyl, imidazolyl, naphthyl, quinolinyl,benzimidazolyl, benzothiofuranyl, fluorenyl, acridinyl, or carbazolyl.

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted C₅₋₆aryl group.

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted phenyl, pyridyl, furanyl, thiofuranyl,indolyl, pyrrolyl, or imidazolyl.

R^(A1) as Optionally Substituted Phenyl

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted phenyl group of the following formula:

wherein each R^(P) is independently a phenyl substituent, and p is aninteger from 0 to 5.

In one embodiment, p is an integer from 0 to 4.

In one embodiment, p is an integer from 0 to 3.

In one embodiment, p is an integer from 0 to 2.

In one embodiment, p is 0 or 1.

R^(A1) as Optionally Substituted Biphenyl

In one embodiment, R^(A1) (and optionally also R^(A2)) is independentlyan optionally substituted biphenyl group of the following formula:

wherein each R^(P) is independently a phenyl substituent, q is aninteger from 0 to 4, and r is an integer from 0 to 5.

In one embodiment, q is an integer from 0 to 3.

In one embodiment, q is an integer from 0 to 2.

In one embodiment, q is 0 or 1.

In one embodiment, q is 0.

In one embodiment, r is an integer from 0 to 4.

In one embodiment, r is an integer from 0 to 3.

In one embodiment, r is an integer from 0 to 2.

In one embodiment, r is 0 or 1.

In one embodiment, r is 0.

In one embodiment, q is 0, and R^(A1) (and optionally also R^(A2)) is anoptionally substituted biphenyl group of the following formula:

R^(A1) as Optionally Substituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is an optionallysubstituted biphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is an optionallysubstituted biphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is an optionallysubstituted biphenyl-4-yl group of the following formula:

R^(A1) as 4′-Substituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

wherein s is an integer from 0 to 4.

In one embodiment, s is an integer from 0 to 3.

In one embodiment, s is an integer from 0 to 2.

In one embodiment, s is 0 or 1.

In one embodiment, s is 0.

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

R^(A1) as 3′-Substituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

R^(A1) as 3′,4′-Disubstituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

wherein t is an integer from 0 to 3.

In one embodiment, t is an integer from 0 to 2.

In one embodiment, t is 0 or 1.

In one embodiment, t is 0.

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

R^(A1) as 2′-Substituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

R^(A1) as 2′,4′-Disubstituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

In one embodiment, R^(A1) (and optionally also R^(A2)) is a substitutedbiphenyl-4-yl group of the following formula:

R^(A1) as Unsubstituted Biphenyl-4-yl

In one embodiment, R^(A1) (and optionally also R^(A2)) is anunsubstituted biphenyl-4-yl group of the following formula:

Some examples of preferred alkane diol derivatives, wherein R^(A2) is—H, are shown below.

Some examples of preferred alkane diol derivatives, wherein R^(A2) is —Hand the alkane diol is 1,4-butanediol, are shown below.

Phenyl Substituents, R^(P)

Examples of phenyl substituents, R^(P), include, but are not limited to,those described below under the heading “substituents.”

Examples of some preferred phenyl substituents, R^(P), include, but arenot limited to, the following:

-   C₁₋₇alkyl (optionally substituted) (including, e.g., unsubstituted    C₁₋₇alkyl, C₁₋₇haloalkyl, C₁₋₇hydroxyalkyl, C₁₋₇carboxyalkyl,    C₁₋₇aminoalkyl, C₅₋₂₀aryl-C₁₋₇alkyl);-   C₃₋₂₀heterocyclyl (optionally substituted);-   C₅₋₂₀aryl group (optionally substituted) (including, e.g.,    C₅₋₂₀carboaryl,-   C₅₋₂₀heteroaryl, C₁₋₇alkyl-C₅₋₂₀aryl and C₅₋₂₀haloaryl); halo;-   hydroxy;-   ether (e.g., C₁₋₇alkoxy);-   acyl (e.g., C₁₋₇alkylacyl, C₅₋₂₀arylacyl);-   carboxy;-   ester;-   acyloxy;-   oxycarboyloxy;-   amido;-   acylamido;-   thioamido;-   tetrazolyl;-   amino;-   nitro;-   cyano;-   sulfhydryl;-   thioether (e.g., C₁₋₇alkylthio);-   sulfonic acid;-   sulfonate; and-   sulfonamido.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   C₁₋₇alkyl (optionally substituted);-   C₃₋₂₀heterocyclyl (optionally substituted);-   C₅₋₂₀aryl group (optionally substituted);-   halo;-   hydroxy;-   ether (e.g., C₁₋₇alkoxy);-   acyl (e.g., C₁₋₇alkylacyl, C₅₋₂₀arylacyl);-   carboxy;-   ester;-   acyloxy;-   amido;-   acylamido;-   amino;-   nitro;-   cyano; and,-   sulfonate.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   -Me, -Et, -iPr, -nPr, -tBu;-   -Ph;-   —F, —Cl, —Br, —I;-   —OH;-   —OMe, —OEt, —O(iPr), —O(nPr), —O(tBu), —OPh, —OBn;-   —C(═O)OH;-   —C(═O)OMe, —C(═O)OEt, —C(═O)O(tBu), —C(═O)OPh;-   —OC(C═O)Me, —OC(C═O)Et, —OC(C═O)(tBu), —OC(C═O)Ph;-   —OC(C═O)OMe, —OC(C═O)OEt, —OC(C═O)O(tBu), —OC(C═O)OPh;-   —C(═O)NH₂, —C(═O)NHMe, —C(═O)NMe₂, —C(═O)NHPh;-   —NHC(═O)Me, —NHC(═O)Et, —NHC(═O)Ph;-   —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂;-   —NO₂;-   —CN; and,-   —S(═O)₂OMe, —S(═O)₂OEt, —S(═O)₂OPh.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   -Me, —F, —Cl, —Br, -1, —OH, —OMe, —NH₂, —NMe₂, —NO₂, and —CN.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   -Me, —F, —Cl, —OH, —OMe, —NH₂, —NMe₂, —NO₂, and —CN.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   —F, —Cl, —Br, -I, —NO₂, and —OH.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   —F, —CI, —Br, and —I, —NO₂.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   —F, —Cl, —Br, —I.

In one embodiment, the phenyl substituents, R^(P), are selected from:

-   —F and —Br.

In one embodiment, the phenyl substituents, R^(P), are —F.

Examples of Some Preferred Fluoro-Substituted Phenyl R^(A1) Groups

Some examples of substituted phenyl groups, suitable as R^(A1) (andoptionally also R^(A2)) include the following:

One especially preferred substituted phenyl group, suitable as R^(A1)(and optionally also R^(A2)) is:

Examples of Some Preferred Substituted Biphenyl-4-yl R^(A1) Groups

Some examples of substituted biphenyl-4-yl groups, suitable as R^(A1)(and optionally also R^(A2)) include the following:

Examples of Some Preferred Fluoro-Substituted Biphenyl-4-yl R^(A1)Groups

Some examples of substituted biphenyl-4-yl groups, suitable as R^(A1)(and optionally also R^(A2)) include the following:

R^(A) as Optionally Substituted Aryl-Alkyl

In one embodiment, R^(A1) alone, R^(A2) alone, or each of R^(A1) andR^(A2), is an optionally substituted C₅₋₂₀aryl-C₁₋₁₇alkyl group.

In one embodiment, R^(A1) alone, R^(A2) alone, or each of R^(A1) andR^(A2), is an optionally substituted C₅₋₆aryl-C₁₋₇alkyl group.

In one embodiment, R^(A1) alone, R^(A2) alone, or each of R^(A1) andR^(A2), is an optionally substituted C₅₋₆aryl-C₁₋₃alkyl group.

Examples of such groups include, but are not limited to, the following:

R^(A) Comprising Optionally Substituted Cycloalkyl

In one embodiment, R^(A1) alone, R^(A2) alone, or each of R^(A1) andR^(A2), is or comprises an optionally substituted C₃₋₇cycloalkyl group.

In one embodiment, R^(A1) alone, R^(A2) alone, or each of R^(A1) andR^(A2), is an optionally substituted C₃₋₇cycloalkyl group or optionallysubstituted C₃₋₇cycloalkyl-C₁₋₇alkyl group.

Examples of such groups include, but are not limited to, the following:

R^(A2) Comprising a Phosphonic Acid Group: Bone Targeting Moieties

In one embodiment, R^(A2) independently is, or comprises, a phosphonicacid group.

In one embodiment, R^(A2) independently is, or comprises, a phosphonicacid group selected from phosphonic acid, and salts (e.g. phosphonates)and esters (e.g., phosphonate esters) thereof.

Without wishing to be bound by any particular theory, it is believedthat such groups act as bone targeting moieties, and improve delivery ofthe compound to the bone envinronment.

Examples of such substituents are shown below. For the phosphonateesters, the groups R¹ and R² are independently C₁₋₇alkyl,C₃₋₂₀heterocyclyl, or C₅₋₂₀aryl, preferably C₁₋₇alkyl.

Where the group is a phosphonate bearing a charge of (−1) or (−2), itwill be associated with a suitable number of cation or cations ofsuitable charge. Examples of suitable cations are discussed below.

Thus, in one embodiment, R^(A2) is, or comprises, a substitutedC₁₋₇alkyl group derived from a bisphosphonate compound.

Examples of bisphosphonate compounds currently in use for the treatmentof osteoporosis, Paget's disease, and cancer associated bone diseaseinclude the following:

Examples of bisphosphonate compounds currently in development includethe following:

In one embodiment, R^(A) independently is, or comprises, a C₁₋₇alkylgroup substituted with one or more groups independently selected fromphosphonic acid, and salts (e.g. phosphonates) and esters (e.g.,phosphonate esters) thereof.

In one embodiment, R^(A2) independently is, or comprises, a C₁₋₇alkylgroup substituted with one group independently selected from phosphonicacid, and salts (e.g. phosphonates) and esters (e.g., phosphonateesters) thereof.

In one embodiment, R^(A2) independently is, or comprises, a C₁₋₇alkylgroup substituted with two groups independently selected from phosphonicacid, and salts (e.g. phosphonates) and esters (e.g., phosphonateesters) thereof.

In one embodiment, R^(A2) independently is, or comprises, a C₁₋₇alkylgroup which is substituted with a bisphosphonic acid group, or a salt orester thereof.

The C₁₋₇alkyl group may optionally be additionally substituted with oneor more other groups.

In one embodiment, R^(A2) independently is, or comprises, a C₁₋₇alkylgroup which comprises a bisphosphonic acid group of the followingformula, or a salt or ester thereof:

In one embodiment, R^(A1), is independently a group selected from groupsof the following formula, or a salt or ester thereof:

In one embodiment, R^(A2), is independently a group of the followingformula, or a salt or ester thereof:

In one embodiment, R^(A2) is independently a group of the followingformula, or a salt or ester thereof, wherein R^(BP) is a bisphosphonatesubstituent, for example, —H, —OH, —Cl, and C₁₋₇alkyl (including, e.g.,unsubstituted C₁₋₇alkyl, C₁₋₇haloalkyl, C₁₋₇hydroxyalkyl,C₁₋₇alkoxyalkyl, C₁₋₇carboxyalkyl, C₁₇aminoalkyl, C₅₋₂₀aryl-C₁₋₇alkyl).

In one embodiment, R^(A2) is independently a group of the followingformula, or a salt or ester thereof:

In one embodiment, R^(A2) is independently a group of the followingformula, or a salt or ester thereof:

R^(A2) Comprising a Ca²⁺ Binding Group: Bone Targeting Moieties

In one embodiment, R^(A2) independently is or comprises a Ca bindinggroup.

The term “Ca²⁺ binding group,” as used herein, pertains to a moietywhich binds (e.g., complexes) one or more Ca²⁺ ions. Without wishing tobe bound by any particular theory, it is believed that such Ca²⁺ bindinggroups act as bone targeting moieties, and improve delivery of thecompound to the bone envinronment.

Examples of Ca²⁺ binding groups include, but are not limited to, thosederived from tetracyclin.

Combination Derivatives of Alkane Diols

In one embodiment, both R¹ and R² are not —H; R¹ and R² are different;and each of R¹ and R² is independently a (non —H) group as describedabove. Such compounds may be conveniently referred to as “asymmetric”compounds.

In one embodiment, both R¹ and R² are not —H; R¹ and R² are different;R¹ is a group of the formula —C(═O)R^(A1), where R^(A1) is an optionallysubstituted aryl group, as described above; and, R² is independently a(non —H) group, as described above.

In one embodiment, both R¹ and R² are not —H; R¹ and R² are different;R¹ is a group of the formula —C(═O)R^(A1), where R^(A1) is an optionallysubstituted aryl group, as described above; and, R² is a group of theformula —C(═O)R^(A2), where R^(A2) is as described above (e.g., aC₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, and isoptionally substituted).

For example, in one embodiment, R^(A1) is an optionally substituted aryland R^(A2) is a C₁₋₇alkyl group. An example of such an embodiment is:

In one embodiment, both R¹ and R² are not —H; R¹ and R² are different;R¹ is a group of the formula —C(═O)R^(A1), where R^(A1) is an optionallysubstituted aryl group, as described above; and, R² is a group of theformula —C(═O)R^(A2), where R^(A2) comprises a bone targeting moiety,for example, phosphonic acid group or a Ca²⁺ binding group, as describedabove.

For example, in one embodiment, the compound has the following formula,or is a salt or ester thereof:

Other Derivatives of Alkane Diols

In one embodiment, the compounds are as described herein, except thatthe group —OR² is replaced with another group.

Thus, one aspect of the present invention pertains to compounds whichhave the following formula:

wherein R¹ and A are as described above, and J is independently selectedfrom hydrogen; halogen; nitrooxy (—ONO₂); ether groups (e.g.,C₁₋₁₇alkoxy); groups which are, or comprise, a phosphonic acid group (asdescribed above); and groups which are, or comprise, a Ca²⁺ bindinggroup (as described above); and pharmaceutically acceptable salts,solvates, amides, esters, ethers, chemically protected forms, orprodrugs thereof.

In one embodiment, the compound has the following formula:

wherein R^(A1), A, and J are as described above.

In one embodiment, J is selected from: hydrogen, halogen, nitrooxy(—ONO₂), and C₁₋₇alkoxy.

In one embodiment, J is selected from: —H, —F, —Cl, —Br, —I, —ONO₂,—OMe, and —OEt.

In one embodiment, J is —H.

In one embodiment, J is selected from: —F, —Cl, —Br, and —I.

In one embodiment, J is —ONO₂.

In one embodiment, J is C₁₋₇alkoxy.

In one embodiment, J is selected from: —OMe and —OEt.

In one embodiment, J is selected from groups which are, or comprise, aphosphonic acid group (as described above).

In one embodiment, J is selected from groups which are, or comprise, aCa²⁺ binding group (as described above).

Examples of Specific Embodiments

Some individual embodiments of the present invention include thefollowing compounds.

1 ABD-0006 4A

2 ABD-0007 4BU

3 ABD-0019 4C

4 ABD-0009 4B

5 ABD-0014 4P

6 ABD-0017 6P

7 ABD-0085 10F

8 ABD-0111 D2,4FB

9 ABD-0096 DBP-4F

10 ABD-0049 4BP-acetate

11 ABD-0008 4MB

12 ABD-0069 4IB

13 ABD-0077 4FB

14 ABD-0106 2,3,6-FB

15 ABD-0107 3,4-FB

16 ABD-0108 2,3,4-FB

17 ABD-0109 2,4,5-FB

18 ABD-0110 2,4-FB

19 ABD-0037 3I

20 ABD-0036 4I

21 ABD-0038 5I

22 ABD-0039 6I

23 ABD-0034 4PT

24 ABD-0059 4BPX

25 ABD-0057 3BP

26 ABD-0056 4BP

27 ABD-0055 5BP

28 ABD-0054 6BP

29 ABD-0095 BP-4F

30 ABD-0070 Me4BP

31 ABD-0072 HO4BP

32 ABD-0089 Xy4BP

33 ABD-0094 Et4BP

34 ABD-0097 4-OMeBP

35 ABD-0098 2-NO₂BP

36 ABD-0099 2-FBP

37 ABD-0100 4-FBP

38 ABD-0102 4-BrBP

39 ABD-0028 4T

40 ABD-0030 5T

41 ABD-0031 6T

42 ABD-0041 3BPA

43 ABD-0042 4BPA

44 ABD-0043 5BPA

45 ABD-0044 6BPA

46 ABD-0032 4N

47 ABD-0033 4H

48 ABD-0035 Bul

49 ABD-0040 BuBPA

50 ABD-0053 BuBP

51 ABD-0090 PBP

52 ABD-0050 4BP-OMe

53 ABD-0086 4BP-Br

54 ABD-0087 4BP-NO₂

55 ABD-0088 4xNO₂-BP

An especially preferred embodiment of the present invention is:

26 ABD-0056 4BP

Another especially preferred embodiment of the present invention is:

7 ABD-0085 10F

Chemical Terms

The term “carbo,” “carbyl,” “hydrocarbon” and “hydrocarbyl,” as usedherein, pertain to compounds and/or groups which have only carbon andhydrogen atoms (but see “carbocyclic” below).

The term “hetero,” as used herein, pertains to compounds and/or groupswhich have at least one heteroatom, for example, multivalent heteroatoms(which are also suitable as ring heteroatoms) such as boron, silicon,nitrogen, phosphorus, oxygen, sulfur, and selenium (more commonlynitrogen, oxygen, and sulfur) and monovalent heteroatoms, such asfluorine, chlorine, bromine, and iodine.

The term “saturated,” as used herein, pertains to compounds and/orgroups which do not have any carbon-carbon double bonds or carbon-carbontriple bonds.

The term “unsaturated,” as used herein, pertains to compounds and/orgroups which have at least one carbon-carbon double bond orcarbon-carbon triple bond.

The term “aliphatic,” as used herein, pertains to compounds and/orgroups which are linear or branched, but not cyclic (also known as“acyclic” or “open-chain” groups).

The term “ring,” as used herein, pertains to a closed ring of from 3 to10 covalently linked atoms, more preferably 3 to 8 covalently linkedatoms, yet more preferably 5 to 6 covalently linked atoms. A ring may bean alicyclic ring or an aromatic ring. The term “alicyclic ring,” asused herein, pertains to a ring which is not an aromatic ring.

The term “carbocyclic ring,” as used herein, pertains to a ring whereinall of the ring atoms are carbon atoms.

The term “heterocyclic ring,” as used herein, pertains to a ring whereinat least one of the ring atoms is a multivalent ring heteroatom, forexample, nitrogen, phosphorus, silicon, oxygen, or sulfur, though morecommonly nitrogen, oxygen, or sulfur. Preferably, the heterocyclic ringhas from 1 to 4 heteroatoms.

The term “cyclic compound,” as used herein, pertains to a compound whichhas at least one ring. The term “cyclyl,” as used herein, pertains to amonovalent moiety obtained by removing a hydrogen atom from a ring atomof a cyclic compound.

Where a cyclic compound has two or more rings, they may be fused (e.g.,as in naphthalene), bridged (e.g., as in norbornane), spiro (e.g., as inspiro[3.3]heptane), or a combination thereof. Cyclic compounds with onering may be referred to as “monocyclic” or “mononuclear,” whereas cycliccompounds with two or more rings may be referred to as “polycyclic” or“polynuclear.”

The term “carbocyclic compound,” as used herein, pertains to a cycliccompound which has only carbocyclic ring(s).

The term “heterocyclic compound,” as used herein, pertains to a cycliccompound which has at least one heterocyclic ring.

The term “aromatic compound,” as used herein, pertains to a cycliccompound which has at least one aromatic ring.

The term “carboaromatic compound,” as used herein, pertains to a cycliccompound which has only carboaromatic ring(s).

The term “heteroaromatic compound,” as used herein, pertains to a cycliccompound which has at least one heteroaromatic ring.

The term “monodentate substituents,” as used herein, pertains tosubstituents which have one point of covalent attachment.

The term “monovalent monodentate substituents,” as used herein, pertainsto substituents which have one point of covalent attachment, via asingle bond. Examples of such substituents include halo, hydroxy, andalkyl.

The term “multivalent monodentate substituents,” as used herein,pertains to substituents which have one point of covalent attachment,but through a double bond or triple bond. Examples of such substituentsinclude oxo, imino, alkylidene, and alklidyne.

The term “bidentate substituents,” as used herein, pertains tosubstituents which have two points of covalent attachment, and which actas a linking group between two other moieties. Examples of suchsubstituents include alkylene and arylene.

Substituents

The phrase “optionally substituted,” as used herein, pertains to aparent group which may be unsubstituted or which may be substituted.

Unless otherwise specified, the term “substituted,” as used herein,pertains to a parent group which bears one or more substitutents. Theterm “substituent” is used herein in the conventional sense and refersto a chemical moiety which is covalently attached to, appended to, or ifappropriate, fused to, a parent group. A wide variety of substituentsare well known, and methods for their formation and introduction into avariety of parent groups are also well known.

The substituents are described in more detail below.

Alkyl: The term “alkyl,” as used herein, pertains to a monovalent moietyobtained by removing a hydrogen atom from a carbon atom of a hydrocarboncompound having from 1 to 20 carbon atoms (unless otherwise specified),which may be aliphatic or alicyclic, and which may be saturated,partially unsaturated, or fully unsaturated. Thus, the term “alkyl”includes the sub-classes alkenyl, alkynyl, cycloalkyl, etc., discussedbelow.

In this context, the prefixes (e.g., C₁₋₄, C₁₋₇, C₁₋₂₀, C₂₋₇, C₃₋₇,etc.) denote the number of carbon atoms, or range of number of carbonatoms. For example, the term “C₁₋₄alkyl,” as used herein, pertains to analkyl group having from 1 to 4 carbon atoms. Examples of groups of alkylgroups include C₁₋₄alkyl (“lower alkyl”), C₁₋₇alkyl, and C₁₋₂₀alkyl.

Examples of (unsubstituted) saturated alkyl groups include, but are notlimited to, methyl (C₁), ethyl (C₂), propyl (C₃), butyl (C₄), pentyl(C₅), hexyl (C₆), heptyl (C₇), octyl (C₈), nonyl (C₉), decyl (C₁₀),n-undecyl (C₁₁), dodecyl (CO₂), tridecyl (CO₃), tetradecyl (C₁₄),pentadecyl (C₁₅), and eicodecyl (C₂₀).

Examples of (unsubstituted) saturated linear alkyl groups include, butare not limited to, methyl (C₁), ethyl (C₂), n-propyl (C₃), n-butyl(C₄), n-pentyl (amyl) (C₅), n-hexyl (C₆), and n-heptyl (C₇).

Examples of (unsubstituted) saturated branched alkyl groups includeiso-propyl (C₃), iso-butyl (C₄), sec-butyl (C₄), tert-butyl (C₄),iso-pentyl (C₅), and neo-pentyl (C₅).

Cycloalkyl: The term “cycloalkyl,” as used herein, pertains to an alkylgroup which is also a cyclyl group; that is, a monovalent moietyobtained by removing a hydrogen atom from an alicyclic ring atom of acyclic hydrocarbon (carbocyclic) compound, which moiety has from 3 to 20ring atoms (unless otherwise specified). Preferably, each ring has from3 to 7 ring atoms.

Examples of (unsubstituted) saturated cylcoalkyl groups include, but arenot limited to, those derived from: cyclopropane (C₃), cyclobutane (C₄),cyclopentane (C₅), cyclohexane (C₆), cycloheptane (C₇), norbornane (C₇),norpinane (C₇), adamantane (C₁₀), and decalin (decahydronaphthalene)(C₁₀).

Examples of (substituted) saturated cycloalkyl groups, which are alsoreferred to herein as “alkyl-cycloalkyl” groups, include, but are notlimited to, methylcyclopropyl, dimethylcyclopropyl, methylcyclobutyl,dimethylcyclobutyl, methylcyclopentyl, dimethylcyclopentyl,methylcyclohexyl, and dimethylcyclohexyl.

Examples of (substituted) unsaturated cyclic alkenyl groups, which arealso referred to herein as “alkyl-cycloalkenyl” groups, include, but arenot limited to, methylcyclopropenyl, dimethylcyclopropenyl,methylcyclobutenyl, dimethylcyclobutenyl, methylcyclopentenyl,dimethylcyclopentenyl, methylcyclohexenyl, and dimethylcyclohexenyl.

Examples of (substituted) cycloalkyl groups, with one or more otherrings fused to the parent cycloalkyl group, include, but are not limitedto, those derived from: indene (C₉), indan (e.g., 2,3-dihydro-1H-indene)(C₉), tetraline (1,2,3,4-tetrahydronaphthalene (C₁₀), fluorene (C₁₃),phenalene (C₁₃). For example, 2H-inden-2-yl is a C₅cycloalkyl group witha substituent (phenyl) fused thereto.

Alkenyl: The term “alkenyl,” as used herein, pertains to an alkyl grouphaving one or more carbon-carbon double bonds. Examples of groups ofalkenyl groups include C₂₋₄alkenyl, C₂₋₇alkenyl, C₂₋₂₀alkenyl.

Examples of (unsubstituted) unsaturated alkenyl groups include, but arenot limited to, ethenyl (vinyl, —CH═CH₂), 1-propenyl (—CH═CH—CH₃),2-propenyl (allyl, —CH—CH═CH₂), isopropenyl (—C(CH₃)═CH₂), butenyl (C₄),pentenyl (C₅), and hexenyl (C₆).

Examples of (unsubstituted) unsaturated cyclic alkenyl groups, which arealso referred to herein as “cycloalkenyl” groups, include, but are notlimited to, cyclopropenyl (C₃), cyclobutenyl (C₄), cyclopentenyl (C₅),and cyclohexenyl (C₆).

Alkynyl: The term “alkynyl,” as used herein, pertains to an alkyl grouphaving one or more carbon-carbon triple bonds. Examples of groups ofalkynyl groups include C₂₋₄alkynyl, C₂₋₇alkynyl, C₂₋₂₀alkynyl.

Examples of (unsubstituted) unsaturated alkynyl groups include, but arenot limited to, ethynyl (ethinyl, —C≡CH) and 2-propynyl (propargyl,—CH₂—C≡CH).

Alkylidene: The term “alkylidene,” as used herein, pertains to adivalent monodentate moiety obtained by removing two hydrogen atoms froma carbon atom of a hydrocarbon compound having from 1 to 20 carbon atoms(unless otherwise specified), which may be aliphatic or alicyclic, or acombination thereof, and which may be saturated, partially unsaturated,or fully unsaturated.

Examples of groups of alkylidene groups include C₁₋₄alkylidene,C₁₋₇alkylidene, C₁₋₂₀alkylidene.

Examples of alkylidene groups include, but are not limited to,methylidene (═CH₂), ethylidene (═CH—CH₃), vinylidene (═C═CH₂), andisopropylidene (═C(CH₃)₂).

Alkylidyne: The term “alkylidyne,” as used herein, pertains to atrivalent monodentate moiety obtained by removing three hydrogen atomsfrom a carbon atom of a hydrocarbon compound having from 1 to 20 carbonatoms (unless otherwise specified), which may be aliphatic or alicyclic,or a combination thereof, and which may be saturated, partiallyunsaturated, or fully unsaturated. Examples of groups of alkylidynegroups include C₁₋₄alkylidyne, C₁₋₇alkylidyne, C₁₋₂₀alkylidyne.

Examples of alkylidyne groups include, but are not limited to,methylidyne (≡CH) and ethylidyne (≡C—CH₃).

Carbocyclyl: The term “carbocyclyl,” as used herein, pertains to amonovalent moiety obtained by removing a hydrogen atom from a ring atomof a carbocyclic compound, which moiety has from 3 to 20 ring atoms(unless otherwise specified). Preferably, each ring has from 3 to 7 ringatoms.

In this context, the prefixes (e.g., C₃₋₂₀, C₃₋₇, C₅₋₆, etc.) denote thenumber of ring atoms, or range of number of ring atoms. For example, theterm “C₅₋₆carbocyclyl,” as used herein, pertains to a carbocyclyl grouphaving 5 or 6 ring atoms. Examples of groups of carbocyclyl groupsinclude C₃₋₂₀carbocyclyl, C₃₋₁₀carbocyclyl, C₅₋₁₀carbocyclyl,C₃₋₇carbocyclyl, and C₅₋₇carbocyclyl.

Examples of carbocyclic groups include, but are not limited to, thosedescribed above as cycloalkyl groups; those described below as carboarylgroups.

Heterocyclyl: The term “heterocyclyl,” as used herein, pertains to amonovalent moiety obtained by removing a hydrogen atom from a ring atomof a heterocyclic compound, which moiety has from 3 to 20 ring atoms(unless otherwise specified), of which from 1 to 10 are ringheteroatoms. Preferably, each ring has from 3 to 7 ring atoms, of whichfrom 1 to 4 are ring heteroatoms.

In this context, the prefixes (e.g., C₃₋₂₀, C₃₋₇, C₅₋₆, etc.) denote thenumber of ring atoms, or range of number of ring atoms, whether carbonatoms or heteroatoms. For example, the term “C₅₋₆heterocyclyl,” as usedherein, pertains to a heterocyclyl group having 5 or 6 ring atoms.Examples of groups of heterocyclyl groups include C₃₋₂₀heterocyclyl,C₃₋₇heterocyclyl, C₅₋₇heterocyclyl.

Examples of (non-aromatic) monocyclic heterocyclyl groups include, butare not limited to, those derived from:

-   N₁: aziridine (C₃), azetidine (C₄), pyrrolidine (tetrahydropyrrole)    (C₅), pyrroline (e.g., 3-pyrroline, 2,5-dihydropyrrole) (C₅),    2H-pyrrole or 3H-pyrrole (isopyrrole, isoazole) (C₅), piperidine    (C₆), dihydropyridine (C₆), tetrahydropyridine (C₆), azepine (C₇);-   O₁: oxirane (C₃), oxetane (C₄), oxolane (tetrahydrofuran) (C₅),    oxole (dihydrofuran) (C₅), oxane (tetrahydropyran) (C₆),    dihydropyran (C₆), pyran (C₆), oxepin (C₇);-   S₁: thiirane (C₃), thietane (C₄), thiolane (tetrahydrothiophene)    (C₅), thiane (tetrahydrothiopyran) (C₆), thiepane (C₇);-   O₂: dioxolane (C₅), dioxane (C₆), and dioxepane (C₇);-   O₃: trioxane (C₆);-   N₂: imidazolidine (C₅), pyrazolidine (diazolidine) (C₅), imidazoline    (C₅), pyrazoline (dihydropyrazole) (C₅), piperazine (C₆);-   N₁O₁: tetrahydrooxazole (C₅), dihydrooxazole (C₅),    tetrahydroisoxazole (C₅), dihydroisoxazole (C₅), morpholine (C₆),    tetrahydrooxazine (C₆), dihydrooxazine (C₆), oxazine (C₆);-   N₁S₁: thiazoline (C₅), thiazolidine (C₅), thiomorpholine (C₆);-   N₂O₁: oxadiazine (C₆);-   O₁S₁: oxathiole (C₅) and oxathiane (thioxane) (C₆); and,-   N₁O₁S₁: oxathiazine (C₆).

Examples of substituted (non-aromatic) monocyclic heterocyclyl groupsinclude saccharides, in cyclic form, for example, furanoses (C₅), suchas arabinofuranose, lyxofuranose, ribofuranose, and xylofuranse, andpyranoses (C₆), such as allopyranose, altropyranose, glucopyranose,mannopyranose, gulopyranose, idopyranose, galactopyranose, andtalopyranose.

Examples of heterocyclyl groups which are also heteroaryl groups aredescribed below with aryl groups.

Aryl: The term “aryl,” as used herein, pertains to a monovalent moietyobtained by removing a hydrogen atom from an aromatic ring atom of anaromatic compound, which moiety has from 3 to 20 ring atoms (unlessotherwise specified). Preferably, each ring has from 5 to 7 ring atoms.

In this context, the prefixes (e.g., C₃₋₂₀, C₅₋₇, C₅₋₆, etc.) denote thenumber of ring atoms, or range of number of ring atoms, whether carbonatoms or heteroatoms. For example, the term “C₅₋₆aryl,” as used herein,pertains to an aryl group having 5 or 6 ring atoms. Examples of groupsof aryl groups include C₃₋₂₀aryl, C₃₋₁₂aryl, C₅₋₁₂aryl, C₅₋₇aryl, andC₅₋₆aryl.

The ring atoms may be all carbon atoms, as in “carboaryl groups” (e.g.,C₅₋₂₀carboaryl).

Examples of carboaryl groups include, but are not limited to, thosederived from benzene (i.e., phenyl) (C₆), naphthalene (C₁₀), azulene(C₁₀), anthracene (C₁₄), phenanthrene (C₁₄), naphthacene (C₁₈), andpyrene (C₁₆).

Examples of aryl groups which comprise fused rings, at least one ofwhich is an aromatic ring, include, but are not limited to, groupsderived from indene (C₉), isoindene (C₉), and fluorene (C₁₃).

Alternatively, the ring atoms may include one or more heteroatoms, as in“heteroaryl groups” (e.g., C₅₋₂₀heteroaryl).

Examples of monocyclic heteroaryl groups include, but are not limitedto, those derived from:

-   N₁: pyrrole (azole) (C₅), pyridine (azine) (C₆);-   O₁: furan (oxole) (C₅);-   S₁: thiophene (thiole) (C₅);-   N₁O₁: oxazole (C₅), isoxazole (C₅), isoxazine (C₆);-   N₂O₁: oxadiazole (furazan) (C₅);-   N₃O₁: oxatriazole (C₅);-   N₁S₁: thiazole (C₅), isothiazole (C₅);-   N₂: imidazole (1,3-diazole) (C₅), pyrazole (1,2-diazole) (C₅),    pyridazine (1,2-diazine) (C₆), pyrimidine (1,3-diazine) (C₆) (e.g.,    cytosine, thymine, uracil), pyrazine (1,4-diazine) (C₆);-   N₃: triazole (C₅), triazine (C₆); and,-   N₄: tetrazole (C₅).

Examples of heterocyclic groups (some of which are also heteroarylgroups) which comprise fused rings, include, but are not limited to:

-   -   C₉heterocyclic groups (with 2 fused rings) derived from        benzofuran (O₁), isobenzofuran (O₁), indole (N₁), isoindole        (N₁), purine (N₄) (e.g., adenine, guanine), benzimidazole (N₂),        benzoxazole (N₁O₁), benzisoxazole (N₁O₁), benzodioxole (O₂),        benzofurazan (N₂O₁), benzotriazole (N₃), benzothiofuran (S₁),        benzothiazole (N₁S₁), benzothiadiazole (N₂S);    -   C₁₀heterocyclic groups (with 2 fused rings) derived from        benzodioxan (O₂), quinoline (N₁), isoquinoline (N₁), benzoxazine        (N₁O₁), benzodiazine (N₂), pyridopyridine (N₂), quinoxaline        (N₂), quinazoline (N₂), phthalazine (N₂), pteridine (N₄);    -   C₁₃heterocyclic groups (with 3 fused rings) derived from        carbazole (N₁), dibenzofuran (O₁), dibenzothiophene (S₁); and,    -   C₁₄heterocyclic groups (with 3 fused rings) derived from        acridine (N₁), xanthene (O₁), phenoxathiin (O₁S₁), phenazine        (N₂), phenoxazine (N₁O₁), phenothiazine (N₁S₁), thianthrene        (S₂), phenanthridine (N₁), phenanthroline (N₂), phenazine (N₂).

Heterocyclic groups (including heteroaryl groups) which have a nitrogenring atom in the form of an —NH— group may be N-substituted, that is, as—NR—. For example, pyrrole may be N-methyl substituted, to giveN-methypyrrole. Examples of N-substitutents include, but are not limitedto C₁₋₇alkyl, C₃₋₂₀heterocyclyl, C₅₋₂₀aryl, and acyl groups.

Heterocyclic groups (including heteroaryl groups) which have a nitrogenring atom in the form of an —N=group may be substituted in the form ofan N-oxide, that is, as —N(→O)=(also denoted —N⁺(→O⁻)=). For example,quinoline may be substituted to give quinoline N-oxide; pyridine to givepyridine N-oxide; benzofurazan to give benzofurazan —N-oxide (also knownas benzofuroxan).

Cyclic groups may additionally bear one or more oxo (═O) groups on ringcarbon atoms. Monocyclic examples of such groups include, but are notlimited to, those derived from:

-   C₅: cyclopentanone, cyclopentenone, cyclopentadienone;-   C₆: cyclohexanone, cyclohexenone, cyclohexadienone;-   O₁: furanone (C₅), pyrone (C₆);-   N₁: pyrrolidone (pyrrolidinone) (C₅), piperidinone (piperidone)    (C₆), piperidinedione (C₆);-   N₂: imidazolidone (imidazolidinone) (C₅), pyrazolone (pyrazolinone)    (C₅), piperazinone (C₆), piperazinedione (C₆), pyridazinone (C₆),    pyrimidinone (C₆) (e.g., cytosine), pyrimidinedione (C₆) (e.g.,    thymine, uracil), barbituric acid (C₆);-   N₁S₁: thiazolone (C₅), isothiazolone (C₅);-   N₁O₁: oxazolinone (C₅).

Polycyclic examples of such groups include, but are not limited to,those derived from:

-   C₉: indenedione;-   C₁₀: tetralone, decalone;-   N₁: oxindole (C₉);-   O₁: benzopyrone (e.g., coumarin, isocoumarin, chromone) (C₁₀);-   N₁O₁: benzoxazolinone (C₉), benzoxazolinone (C₁₀);-   N₂: quinazolinedione (C₁₀);-   N₄: purinone (C₉) (e.g., guanine).

Still more examples of cyclic groups which bear one or more oxo (═O)groups on ring carbon atoms include, but are not limited to, thosederived from:

-   -   cyclic anhydrides (—C(═O)—O—C(═O)— in a ring), including but not        limited to maleic anhydride (C₅), succinic anhydride (C₅), and        glutaric anhydride (C₆);    -   cyclic carbonates (—O—C(═O)—O— in a ring), such as ethylene        carbonate (C₅) and 1,2-propylene carbonate (C₅);    -   imides (—C(═O)—NR—C(═O)— in a ring), including but not limited        to, succinimide (C₅), maleimide (C₅), phthalimide, and        glutarimide (C₆);    -   lactones (cyclic esters, —O—C(═O)— in a ring), including, but        not limited to, β-propiolactone, γ-butyrolactone,        δ-valerolactone (2-piperidone), and ε-caprolactone;    -   lactams (cyclic amides, —NR—C(═O)— in a ring), including, but        not limited to, β-propiolactam (C₄), γ-butyrolactam        (2-pyrrolidone) (C₅), δ-valerolactam (C₆), and ε-caprolactam        (C₇);    -   cyclic carbamates (—O—C(═O)—NR— in a ring), such as        2-oxazolidone (C₅);    -   cyclic ureas (—NR—C(═O)—NR— in a ring), such as 2-imidazolidone        (C₅) and pyrimidine-2,4-dione (e.g., thymine, uracil) (C₆).

The above alkyl, alkylidene, alkylidyne, heterocyclyl, and aryl groups,whether alone or part of another substituent, may themselves optionallybe substituted with one or more groups selected from themselves and theadditional substituents listed below.

Hydrogen: —H. Note that if the substituent at a particular position ishydrogen, it may be convenient to refer to the compound as being“unsubstituted” at that position.

Halo: —F, —Cl, —Br, and —I.

Hydroxy: —OH.

Ether: —OR, wherein R is an ether substituent, for example, a C₁₋₇alkylgroup (also referred to as a C₁₋₇alkoxy group, discussed below), aC₃₋₂₀heterocyclyl group (also referred to as a C₃₋₂₀heterocyclyloxygroup), or a C₅₋₂₀aryl group (also referred to as a C₅₋₂₀aryloxy group),preferably a C₁₋₇alkyl group.

C₁₋₇alkoxy: —OR, wherein R is a C₁₋₇alkyl group. Examples of C₁₋₇alkoxygroups include, but are not limited to, —OMe (methoxy), —OEt (ethoxy),—O(nPr) (n-propoxy), —O(iPr) (isopropoxy), —O(nBu) (n-butoxy), —O(sBu)(sec-butoxy), —O(iBu) (isobutoxy), and —O(tBu) (tert-butoxy).

Acetal: —CH(OR¹)(OR²), wherein R¹ and R² are independently acetalsubstituents, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group,or a C₅₋₂₀aryl group, preferably a C₁₋₇alkyl group, or, in the case of a“cyclic” acetal group, R¹ and R², taken together with the two oxygenatoms to which they are attached, and the carbon atoms to which they areattached, form a heterocyclic ring having from 4 to 8 ring atoms.Examples of acetal groups include, but are not limited to, —CH(OMe)₂,—CH(OEt)₂, and —CH(OMe)(OEt).

Hemiacetal: —CH(OH)(OR¹), wherein R¹ is a hemiacetal substituent, forexample, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably a C₁₋₇alkyl group. Examples of hemiacetal groupsinclude, but are not limited to, —CH(OH)(OMe) and —CH(OH)(OEt).

Ketal: —CR(OR¹)(OR²), where R¹ and R² are as defined for acetals, and Ris a ketal substituent other than hydrogen, for example, a C₁₋₇alkylgroup, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably aC₁₋₇alkyl group.

Examples ketal groups include, but are not limited to, —C(Me)(OMe)₂,—C(Me)(OEt)₂, —C(Me)(OMe)(OEt), —C(Et)(OMe)₂, —C(Et)(OEt)₂, and—C(Et)(OMe)(OEt).

Hemiketal: —CR(OH)(OR¹), where R¹ is as defined for hemiacetals, and Ris a hemiketal substituent other than hydrogen, for example, a C₁₋₇alkylgroup, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably aC₁₋₇alkyl group. Examples of hemiacetal groups include, but are notlimited to, —C(Me)(OH)(OMe), —C(Et)(OH)(OMe), —C(Me)(OH)(OEt), and—C(Et)(OH)(OEt).

Oxo (keto, -one): ═O.

Thione (thioketone): ═S.

Imino (imine): —NR, wherein R is an imino substituent, for example,hydrogen, C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably hydrogen or a C₁₋₇alkyl group. Examples of estergroups include, but are not limited to, ═NH, ═NMe, ═NEt, and ═NPh.

Formyl (carbaldehyde, carboxaldehyde): —C(═O)H.

Acyl (keto): —C(═O)R, wherein R is an acyl substituent, for example, aC₁₋₇alkyl group (also referred to as C₁₋₇alkylacyl or C₁₋₇alkanoyl), aC₃₋₂₀heterocyclyl group (also referred to as C₃₋₂₀heterocyclylacyl), ora C₅₋₂₀aryl group (also referred to as C₅₋₂₀arylacyl), preferably aC₁₋₇alkyl group. Examples of acyl groups include, but are not limitedto, —C(═O)CH₃ (acetyl), —C(═O)CH₂CH₃ (propionyl), —C(═O)C(CH₃)₃(t-butyryl), and —C(═O)Ph (benzoyl, phenone).

Carboxy (carboxylic acid): —C(═O)OH.

Thiocarboxy (thiocarboxylic acid): —C(═S)SH.

Thiolocarboxy (thiolocarboxylic acid): —C(═O)SH.

Thionocarboxy (thionocarboxylic acid): —C(═S)OH.

Imidic acid: —C(═NH)OH.

Hydroxamic acid: —C(═NOH)OH.

Ester (carboxylate, carboxylic acid ester, oxycarbonyl): —C(═O)OR,wherein R is an ester substituent, for example, a C₁₋₇alkyl group, aC₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably a C₁₋₇alkylgroup. Examples of ester groups include, but are not limited to,—C(═O)OCH₃, —C(═O)OCH₂CH₃, —C(═O)OC(CH₃)₃, and —C(═O)OPh.

Acyloxy (reverse ester): —OC(═O)R, wherein R is an acyloxy substituent,for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or aC₅₋₂₀aryl group, preferably a C₁₋₇alkyl group. Examples of acyloxygroups include, but are not limited to, —OC(═O)CH₃ (acetoxy),—OC(═O)CH₂CH₃, —OC(═O)C(CH₃)₃, —OC(═O)Ph, and —OC(═O)CH₂Ph.

Oxycarboyloxy: —OC(═O)OR, wherein R is an ester substituent, forexample, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably a C₁₋₇alkyl group. Examples of ester groups include,but are not limited to, —OC(═O)OCH₃, —OC(═O)OCH₂CH₃, —OC(═O)OC(CH₃)₃,and —OC(═O)OPh.

Amido (carbamoyl, carbamyl, aminocarbonyl, carboxamide): —C(═O)NR¹R²,wherein R¹ and R² are independently amino substituents, as defined foramino groups. Examples of amido groups include, but are not limited to,—C(═O)NH₂, —C(═O)NHCH₃, —C(═O)N(CH₃)₂, —C(═O)NHCH₂CH₃, and—C(═O)N(CH₂CH₃)₂, as well as amido groups in which R¹ and R², togetherwith the nitrogen atom to which they are attached, form a heterocyclicstructure as in, for example, piperidinocarbonyl, morpholinocarbonyl,thiomorpholinocarbonyl, and piperazinocarbonyl.

Acylamido (acylamino): —NR¹C(═O)R², wherein R¹ is an amide substituent,for example, hydrogen, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, ora C₅₋₂₀aryl group, preferably hydrogen or a C₁₋₇alkyl group, and R² isan acyl substituent, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclylgroup, or a C₅₋₂₀aryl group, preferably hydrogen or a C₁₋₇alkyl group.Examples of acylamide groups include, but are not limited to,—NHC(═O)CH₃, —NHC(═O)CH₂CH₃, and —NHC(═O)Ph. R¹ and R² may together forma cyclic structure, as in, for example, succinimidyl, maleimidyl, andphthalimidyl:

Thioamido (thiocarbamyl): —C(═S)NR¹R², wherein R¹ and R² areindependently amino substituents, as defined for amino groups. Examplesof amido groups include, but are not limited to, —C(═S)NH₂, —C(═S)NHCH₃,—C(═S)N(CH₃)₂, and —C(═S)NHCH₂CH₃.

Ureido: —N(R¹)CONR²R³ wherein R² and R³ are independently aminosubstituents, as defined for amino groups, and R1 is a ureidosubstituent, for example, hydrogen, a C₁₋₇alkyl group, aC₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably hydrogen or aC₁₋₇alkyl group. Examples of ureido groups include, but are not limitedto, —NHCONH₂, —NHCONHMe, —NHCONHEt, —NHCONMe₂, —NHCONEt₂, —NMeCONH₂,—NMeCONHMe, —NMeCONHEt, —NMeCONMe₂, and —NMeCONEt₂.

Guanidino: —NH—C(═NH)NH₂.

Tetrazolyl: a five membered aromatic ring having four nitrogen atoms andone carbon atom,

Amino: —NR¹R², wherein R¹ and R² are independently amino substituents,for example, hydrogen, a C₁₋₇alkyl group (also referred to asC₁₋₇alkylamino or di-C₁₋₇alkylamino), a C₃₋₂₀heterocyclyl group, or aC₅₋₂₀aryl group, preferably H or a 12 C₁₋₇alkyl group, or, in the caseof a “cyclic” amino group, R¹ and R², taken together with the nitrogenatom to which they are attached, form a heterocyclic ring having from 4to 8 ring atoms. Amino groups may be primary (—NH₂), secondary (—NHR¹),or tertiary (—NHR¹R²), and in cationic form, may be quaternary(—⁺NR¹R²R³). Examples of amino groups include, but are not limited to,—NH₂, —NHCH₃, —NHC(CH₃)₂, —N(CH₃)₂, —N(CH₂CH₃)₂, and —NHPh. Examples ofcyclic amino groups include, but are not limited to, aziridino,azetidino, pyrrolidino, piperidino, piperazino, morpholino, andthiomorpholino.

Amidine (amidino): —C(═NR)NR₂, wherein each R is an amidine substituent,for example, hydrogen, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, ora C₅₋₂₀aryl group, preferably H or a C₁₋₇alkyl group. Examples ofamidine groups include, but are not limited to, —C(═NH)NH₂, —C(═NH)NMe₂,and —C(═NMe)NMe₂.

Nitro: —NO₂.

Nitroso: —NO.

Azido: —N₃.

Cyano (nitrile, carbonitrile): —CN.

Isocyano: —NC.

Cyanato: —OCN.

Isocyanato: —NCO.

Thiocyano (thiocyanato): —SCN.

Isothiocyano (isothiocyanato): —NCS.

Sulfhydryl (thiol, mercapto): —SH.

Thioether (sulfide): —SR, wherein R is a thioether substituent, forexample, a C₁₋₇alkyl group (also referred to as a C₁₋₇alkylthio group),a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably a C₁₋₇alkylgroup. Examples of C₁₋₇alkylthio groups include, but are not limited to,—SCH₃ and —SCH₂CH₃.

Disulfide: —SS—R, wherein R is a disulfide substituent, for example, aC₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group,preferably a C₁₋₇alkyl group (also referred to herein as C₁₋₇alkyldisulfide). Examples of C₁₋₇alkyl disulfide groups include, but are notlimited to, —SSCH₃ and —SSCH₂CH₃.

Sulfonic acid (sulfo): —S(═O)₂OH, —SO₃H.

Sulfonate (sulfonic acid ester): —S(═O)₂OR, wherein R is a sulfonatesubstituent, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group,or a C₅₋₂₀aryl group, preferably a C₁₋₇alkyl group. Examples ofsulfonate groups include, but are not limited to, —S(═O)₂OCH₃ and—S(═O)₂OCH₂CH₃.

Sulfinic acid: —S(═O)OH, —SO₂H.

Sulfinate (sulfinic acid ester): —S(═O)OR; wherein R is a sulfinatesubstituent, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group,or a C₅₋₂₀aryl group, preferably a C₁₋₇alkyl group. Examples ofsulfinate groups include, but are not limited to, —S(═O)OCH₃ and—S(═O)OCH₂CH₃.

Sulfate: —OS(═O)₂OR; wherein R is a sulfate substituent, for example, aC₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group,preferably a C₁₋₇alkyl group. Examples of sulfate groups include, butare not limited to, —OS(═O)₂OCH₃ and —SO(═O)₂OCH₂CH₃.

Sulfone (sulfonyl): —S(═O)₂R, wherein R is a sulfone substituent, forexample, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably a C₁₋₇alkyl group, for example, a fluorinated orperfluorinated C₁₋₇alkyl group. Examples of sulfone groups include, butare not limited to, —S(═O)₂CH₃ (methanesulfonyl, mesyl), —S(═O)₂CF₃(triflyl), —S(═O)₂CH₂CH₃ (esyl), —S(═O)₂C₄F₉ (nonaflyl), —S(═O)₂CH₂CF₃(tresyl), —S(═O)₂Ph (phenylsulfonyl, besyl), 4-methylphenylsulfonyl(tosyl), 4-chlorophenylsulfonyl (closyl), 4-bromophenylsulfonyl(brosyl), 4-nitrophenyl (nosyl), 2-naphthalenesulfonate (napsyl), and5-dimethylamino-naphthalen-1-ylsulfonate (dansyl).

Sulfine (sulfinyl, sulfoxide): —S(═O)R, wherein R is a sulfinesubstituent, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group,or a C₅₋₂₀aryl group, preferably a C₁₋₇alkyl group. Examples of sulfinegroups include, but are not limited to, —S(═O)CH₃ and —S(═O)CH₂CH₃.

Sulfonyloxy: —OS(═O)₂R, wherein R is a sulfonyloxy substituent, forexample, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably a C₁₋₁₇alkyl group. Examples of sulfonyloxy groupsinclude, but are not limited to, —OS(═O)₂CH₃ (mesylate) and—OS(═O)₂CH₂CH₃ (esylate).

Sulfinyloxy: —OS(═O)R, wherein R is a sulfinyloxy substituent, forexample, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀arylgroup, preferably a C₁₋₇alkyl group. Examples of sulfinyloxy groupsinclude, but are not limited to, —OS(═O)CH₃ and —OS(═O)CH₂CH₃.

Sulfamino: —NR¹S(═O)₂OH, wherein R¹ is an amino substituent, as definedfor amino groups. Examples of sulfamino groups include, but are notlimited to, —NHS(═O)₂OH and —N(CH₃)S(═O)₂OH.

Sulfonamino: —NR¹S(═O)₂R, wherein R¹ is an amino substituent, as definedfor amino groups, and R is a sulfonamino substituent, for example, aC₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group,preferably a C₁₋₇alkyl group. Examples of sulfonamino groups include,but are not limited to, —NHS(═O)₂CH₃ and —N(CH₃)S(═O)₂C₆H₅.

Sulfinamino: —NR¹S(═O)R, wherein R¹ is an amino substituent, as definedfor amino groups, and R is a sulfinamino substituent, for example, aC₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group,preferably a C₁₋₇alkyl group.

Examples of sulfinamino groups include, but are not limited to,—NHS(═O)CH₃ and —N(CH₃)S(═O)C₆H₅.

Sulfamyl: —S(═O)NR¹R², wherein R¹ and R² are independently aminosubstituents, as defined for amino groups. Examples of sulfamyl groupsinclude, but are not limited to, —S(═O)NH₂, —S(═O)NH(CH₃),—S(═O)N(CH₃)₂, —S(═O)NH(CH₂CH₃), —S(═O)N(CH₂CH₃)₂, and —S(═O)NHPh.

Sulfonamido: —S(═O)₂NR¹R² ₁ wherein R¹ and R² are independently aminosubstituents, as defined for amino groups. Examples of sulfonamidogroups include, but are not limited to, —S(═O)₂NH₂, —S(═O)₂NH(CH₃),—S(═O)₂N(CH₃)₂, —S(═O)₂NH(CH₂CH₃), —S(═O)₂N(CH₂CH₃)₂, and —S(═O)₂NHPh.

Phosphino (phosphine): —PR₂, wherein R is a phosphino substituent, forexample, —H, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or aC₅₋₂₀aryl group, preferably —H, a C₁₋₇alkyl group, or a C₅₋₂₀aryl group.Examples of phosphino groups include, but are not limited to, —PH₂,—P(CH₃)₂, —P(CH₂CH₃)₂, —P(t-Bu)₂, and —P(Ph)₂.

Phospho: —P(═O)₂.

Phosphinyl (phosphine oxide): —P(═O)R₂, wherein R is a phosphinylsubstituent, for example, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group,or a C₅₋₂₀aryl group, preferably a C₁₋₇alkyl group or a C₅₋₂₀aryl group.Examples of phosphinyl groups include, but are not limited to,—P(═O)(CH₃)₂, —P(═O)(CH₂CH₃)₂, —P(═O)(t-Bu)₂, and —P(═O)(Ph)₂.

Phosphonic acid (phosphono): —P(═O)(OH)₂.

Phosphonate (phosphono ester): —P(═O)(OR)₂, where R is a phosphonatesubstituent, for example, —H, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclylgroup, or a C₅₋₂₀aryl group, preferably —H, a C₁₋₇alkyl group, or aC₅₋₂₀aryl group. Examples of phosphonate groups include, but are notlimited to, —P(═O)(OCH₃)₂, —P(═O)(OCH₂CH₃)₂, —P(═O)(O-t-Bu)₂, and—P(═O)(OPh)₂.

Phosphoric acid (phosphonooxy): —OP(═O)(OH)₂.

Phosphate (phosphonooxy ester): —OP(═O)(OR)₂, where R is a phosphatesubstituent, for example, —H, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclylgroup, or a C₅₋₂₀aryl group, preferably —H, a C₁₋₇alkyl group, or aC₅₋₂₀aryl group. Examples of phosphate groups include, but are notlimited to, —OP(═O)(OCH₃)₂, —OP(═O)(OCH₂CH₃)₂, —OP(═O)(O-t-Bu)₂, and—OP(═O)(OPh)₂.

Phosphorous acid: —OP(OH)₂.

Phosphite: —OP(OR)₂, where R is a phosphite substituent, for example,—H, a C₁₋₇alkyl group, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group,preferably —H, a C₁₋₇alkyl group, or a C₅₋₂₀aryl group. Examples ofphosphite groups include, but are not limited to, —OP(OCH₃)₂,—OP(OCH₂CH₃)₂, —OP(O-t-Bu)₂, and —OP(OPh)₂.

Phosphoramidite: —OP(OR¹)—NR² ₂, where R¹ and R² are phosphoramiditesubstituents, for example, —H, a (optionally substituted) C₁₋₇alkylgroup, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably —H, aC₁₋₇alkyl group, or a C₅₋₂₀aryl group. Examples of phosphoramiditegroups include, but are not limited to, —OP(OCH₂CH₃)—N(CH₃)₂,—OP(OCH₂CH₃)—N(i-Pr)₂, and —OP(OCH₂CH₂CN)—N(i-Pr)₂.

Phosphoramidate: —OP(═O)(OR¹)—NR² ₂, where R¹ and R² are phosphoramidatesubstituents, for example, —H, a (optionally substituted) C₁₋₇alkylgroup, a C₃₋₂₀heterocyclyl group, or a C₅₋₂₀aryl group, preferably —H, aC₁₋₇alkyl group, or a C₅₋₂₀aryl group. Examples of phosphoramidategroups include, but are not limited to, —OP(═O)(OCH₂CH₃)—N(CH₃)₂,—OP(═O)(OCH₂CH₃)—N(i-Pr)₂, and —OP(═O)(OCH₂CH₂CN)—N(i-Pr)₂.

In many cases, substituents may themselves be substituted. For example,a C₁₋₇alkyl group may be substituted with, for example, hydroxy (alsoreferred to as a C₁₋₇hydroxyalkyl group), C₁₋₇alkoxy (also referred toas a C₁₋₇alkoxyalkyl group), amino (also referred to as a C₁₋₇aminoalkylgroup), halo (also referred to as a C₁₋₇haloalkyl group), carboxy (alsoreferred to as a C₁₋₇carboxyalkyl group), and C₅₋₂₀aryl (also referredto as a C₅₋₂₀aryl-C₁₋₇alkyl group).

Similarly, a C₅₋₂₀aryl group may be substituted with, for example,hydroxy (also referred to as a C₅₋₂₀hydroxyaryl group), halo (alsoreferred to as a C₅₋₂₀haloaryl group), amino (also referred to as aC₅₋₂₀aminoaryl group, e.g., as in aniline), C₁₋₇alkyl (also referred toas a C₁₋₇alkyl-C₅₋₂₀aryl group, e.g., as in toluene), and C₁₋₇alkoxy(also referred to as a C₁₋₇alkoxy-C₅₋₂₀aryl group, e.g., as in anisole).

These and other specific examples of such substituted-substituents aredescribed below.

C₁₋₇haloalkyl group: The term “C₁₋₇haloalkyl group,” as used herein,pertains to a C₁₋₇alkyl group in which at least one hydrogen atom (e.g.,1, 2, 3) has been replaced with a halogen atom (e.g., F, Cl, Br, I). Ifmore than one hydrogen atom has been replaced with a halogen atom, thehalogen atoms may independently be the same or different. Every hydrogenatom may be replaced with a halogen atom, in which case the group mayconveniently be referred to as a C₁₋₇perhaloalkyl group.” Examples ofC₁₋₇haloalkyl groups include, but are not limited to, —CF₃, —CHF₂,—CH₂F, —CCl₃, —CBr₃, —CH₂CH₂F, —CH₂CHF₂, and —CH₂CF₃.

C₁₋₇haloalkoxy: —OR, wherein R is a C₁₋₇haloalkyl group. Examples ofC₁₋₇haloalkoxy groups include, but are not limited to, —OCF₃, —OCHF₂,—OCH₂F, —OCCl₃, —OCBr₃, —OCH₂CH₂F, —OCH₂CHF₂, and —OCH₂CF₃.

C₁₋₇hydroxyalkyl: The term “C₁₋₇hydroxyalkyl group,” as used herein,pertains to a C₁₋₇alkyl group in which at least one hydrogen atom hasbeen replaced with a hydroxy group. Examples of C₁₋₇hydroxyalkyl groupsinclude, but are not limited to, —CH₂OH, —CH₂CH₂OH, and —CH(OH)CH₂OH.

C₁₋₇carboxyalkyl: The term “C₁₋₇carboxyalkyl group,” as used herein,pertains to a C₁₋₇alkyl group in which at least one hydrogen atom hasbeen replaced with a carboxy group. Examples of C₁₋₇carboxyalkyl groupsinclude, but are not limited to, —CH₂COOH and —CH₂CH₂COOH.

C₁₋₇aminoalkyl: The term “C₁₋₇aminoalkyl group,” as used herein,pertains to a C₁₋₇alkyl group in which at least one hydrogen atom hasbeen replaced with an amino group. Examples of C₁₋₇aminoalkyl groupsinclude, but are not limited to, —CH₂NH₂, —CH₂CH₂NH₂, and—CH₂CH₂N(CH₃)₂.

C₁₋₇aminoalkylamino: The term “C₁₋₇aminoalkylamino,” as used herein,pertains to an amino group, —NR¹R², in which one of the substituents, R¹or R², is itself a C₁₋₇aminoalkyl group (—C₁₋₇alkyl-NR¹R²). TheC₁₇aminoalkylamino may be represented, for example, by the formula—NR¹-C₁₋₇alkyl-NR¹R². Examples of amino-C₁₋₇alkylamino groups include,but are not limited to, groups of the formula —NR¹(CH₂)_(n)NR¹R², wheren is 1 to 6, for example, —NHCH₂NH₂, —NH(CH₂)₂NH₂, —NH(CH₂)₃NH₂,—NH(CH₂)₄NH₂, —NH(CH₂)₅NH₂, —NH(CH₂)₆NH₂, —NHCH₂NH(Me), —NH(CH₂)₂NH(Me),—NH(CH₂)₃NH(Me), —NH(CH₂)₄NH(Me), —NH(CH₂)₅NH(Me), —NH(CH₂)₆NH(Me),—NHCH₂NH(Et), —NH(CH₂)₂NH(Et), —NH(CH₂)₃NH(Et), —NH(CH₂)₄NH(Et),—NH(CH₂)₅NH(Et), and —NH(CH₂)₆NH(Et).

C₃₋₇cycloalkyl-C₁₋₇alkyl: The term “,” as used herein, describes certainC₁₋₇alkyl groups which have been substituted with a C₃₋₇cycloalkylgroup. Examples of such groiups include, but are not limited to,cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, andcyclohexylmethyl.

C₃₋₇cycloalkenyl-C₁₋₇alkyl: The term “,” as used herein, describescertain C₁₋₇alkyl groups which have been substituted with aC₃₋₇cycloalkenyl group.

Examples of such groiups include, but are not limited to,cyclopropenylmethyl and cyclohexenylmethyl.

C₁₋₇alkyl-C₅₋₂₀aryl: The term “C₁₋₇alkyl-C₅₋₂₀aryl,” as used herein,describes certain C₅₋₂₀aryl groups which have been substituted with aC₁₋₇alkyl group. Examples of such groups include, but are not limitedto, tolyl (as in toluene), xylyl (as in xylene), mesityl (as inmesitylene), styryl (as in styrene), and cumenyl (as in cumene).

C₁₋₇alkyl-C₅₋₂₀aryloxy: The term “C₁₋₇alkyl-C₅₋₂₀aryloxy,” as usedherein, describes certain C₅₋₂₀aryloxy groups which have beensubstituted with a C₁₋₇alkyl group. Examples of such groups include, butare not limited to, tolyloxy, xylyloxy, mesityloxy, and cumenyloxy.

C₅₋₂₀aryl-C₁₋₇alkyl: The term “C₅₋₂₀aryl-C₁₋₇alkyl,” as used herein,describers certain C₁₋₇alkyl groups which have been substituted with aC₅₋₂₀aryl group. Examples of such groups include, but are not limitedto, benzyl (phenylmethyl), tolylmethyl, phenylethyl, triphenylmethyl(trityl), and cinnamyl (3-phenyl-2-propenyl, C₆H₅—CH═CH—CH₂—).

C₅₋₂₀aryl-C₁₋₇alkoxy: The term “C₅₋₂₀aryl-C₁₋₇alkoxy,” as used herein,describes certain C₁₋₇alkoxy groups which have been substituted with aC₅₋₂₀aryl group. Examples of such groups include, but are not limitedto, benzyloxy, tolylmethoxy, and phenylethoxy.

C₅₋₂₀haloaryl: The term “C₅₋₂₀haloaryl,” as used herein, describescertain C₅₋₂₀aryl groups which have been substituted with one or morehalo groups. Examples of such groups include, but are not limited to,halophenyl (e.g., fluorophenyl, chlorophenyl, bromophenyl, oriodophenyl, whether ortho-, meta-, or para-substituted), dihalophenyl,trihalophenyl, tetrahalophenyl, and pentahalophenyl.

Includes Other Forms

Unless otherwise specified, included in the above are the well knownionic, salt, solvate, and protected forms of these substituents. Forexample, a reference to carboxylic acid (—COOH) also includes theanionic (carboxylate) form (—COO⁻), a salt or solvate thereof, as wellas conventional protected forms. Similarly, a reference to an aminogroup includes the protonated form (—N⁺HR¹R²), a salt or solvate of theamino group, for example, a hydrochloride salt, as well as conventionalprotected forms of an amino group. Similarly, a reference to a hydroxylgroup also includes the anionic form (—O⁻), a salt or solvate thereof,as well as conventional protected forms of a hydroxyl group.

Isomers, Salts, Solvates, Protected Forms, and Prodrugs

Certain compounds may exist in one or more particular geometric,optical, enantiomeric, diasteriomeric, epimeric, stereoisomeric,tautomeric, conformational, or anomeric forms, including but not limitedto, cis- and trans-forms; E- and Z-forms; c-, t-, and r-forms; endo- andexo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+)and (−) forms; keto-, enol-, and enolate-forms; syn- and anti-forms;synclinal- and anticlinal-forms; α- and β-forms; axial and equatorialforms; boat-, chair-, twist-, envelope-, and halfchair-forms; andcombinations thereof, hereinafter collectively referred to as “isomers”(or “isomeric forms”).

Note that, except as discussed below for tautomeric forms, specificallyexcluded from the term “isomers,” as used herein, are structural (orconstitutional) isomers (i.e., isomers which differ in the connectionsbetween atoms rather than merely by the position of atoms in space). Forexample, a reference to a methoxy group, —OCH₃, is not to be construedas a reference to its structural isomer, a hydroxymethyl group, —CH₂OH.Similarly, a reference to ortho-chlorophenyl is not to be construed as areference to its structural isomer, meta-chlorophenyl. However, areference to a class of structures may well include structurallyisomeric forms falling within that class (e.g., C₁₋₇alkyl includesn-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl;methoxyphenyl includes ortho-, meta-, and para-methoxyphenyl).

The above exclusion does not pertain to tautomeric forms, for example,keto-, enol-, and enolate-forms, as in, for example, the followingtautomeric pairs: keto/enol (illustrated below), imine/enamine,amide/imino alcohol, amidinelamidine, nitroso/oxime,thioketone/enethiol, N-nitroso/hyroxyazo, and nitro/aci-nitro.

Note that specifically included in the term “isomer” are compounds withone or more isotopic substitutions. For example, H may be in anyisotopic form, including ¹H, ²H (D), and ³H (T); C may be in anyisotopic form, including ¹²C, ¹³C, and ¹⁴C; 0 may be in any isotopicform, including ¹⁶O and ¹⁸O; and the like.

Unless otherwise specified, a reference to a particular compoundincludes all such isomeric forms, including (wholly or partially)racemic and other mixtures thereof. Methods for the preparation (e.g.,asymmetric synthesis) and separation (e.g., fractional crystallisationand chromatographic means) of such isomeric forms are either known inthe art or are readily obtained by adapting the methods taught herein,or known methods, in a known manner.

Unless otherwise specified, a reference to a particular compound alsoincludes ionic, salt, solvate, and protected forms of thereof, forexample, as discussed below.

It may be convenient or desirable to prepare, purify, and/or handle acorresponding salt of the active compound, for example, apharmaceutically-acceptable salt. Examples of pharmaceuticallyacceptable salts are discussed in Berge et al., 1977, “PharmaceuticallyAcceptable Salts,” J. Pharm. Sci., Vol. 66, pp. 1-19.

For example, if the compound is anionic, or has a functional group whichmay be anionic (e.g., —COOH may be —COO⁻), then a salt may be formedwith a suitable cation. Examples of suitable inorganic cations include,but are not limited to, alkali metal ions such as Na⁺ and K⁺, alkalineearth cations such as Ca²⁺ and Mg²⁺, and other cations such as Al⁺³.Examples of suitable organic cations include, but are not limited to,ammonium ion (i.e., NH₄ ⁺) and substituted ammonium ions (e.g., NH₃R⁺,NH₂R₂ ⁺, NHR₃ ⁺, NR₄ ⁺). Examples of some suitable substituted ammoniumions are those derived from: ethylamine, diethylamine,dicyclohexylamine, triethylamine, butylamine, ethylenediamine,ethanolamine, diethanolamine, piperazine, benzylamine,phenylbenzylamine, choline, meglumine, and tromethamine, as well asamino acids, such as lysine and arginine. An example of a commonquaternary ammonium ion is N(CH₃)₄ ⁺.

If the compound is cationic, or has a functional group which may becationic (e.g., —NH₂ may be —NH₃ ⁺), then a salt may be formed with asuitable anion. Examples of suitable inorganic anions include, but arenot limited to, those derived from the following inorganic acids:hydrochloric, hydrobromic, hydroiodic, sulfuric, sulfurous, nitric,nitrous, phosphoric, and phosphorous.

Examples of suitable organic anions include, but are not limited to,those derived from the following organic acids: 2-acetyoxybenzoic,acetic, ascorbic, aspartic, benzoic, camphorsulfonic, cinnamic, citric,edetic, ethanedisulfonic, ethanesulfonic, fumaric, glucheptonic,gluconic, glutamic, glycolic, hydroxymaleic, hydroxynaphthalenecarboxylic, isethionic, lactic, lactobionic, lauric, maleic, malic,methanesulfonic, mucic, oleic, oxalic, palmitic, pamoic, pantothenic,phenylacetic, phenylsulfonic, propionic, pyruvic, salicylic, stearic,succinic, sulfanilic, tartaric, toluenesulfonic, and valeric. Examplesof suitable polymeric organic anions include, but are not limited to,those derived from the following polymeric acids: tannic acid,carboxymethyl cellulose.

It may be convenient or desirable to prepare, purify, and/or handle acorresponding solvate of the active compound. The term “solvate” is usedherein in the conventional sense to refer to a complex of solute (e.g.,active compound, salt of active compound) and solvent. If the solvent iswater, the solvate may be conveniently referred to as a hydrate, forexample, a mono-hydrate, a di-hydrate, a tri-hydrate, etc.

It may be convenient or desirable to prepare, purify, and/or handle theactive compound in a chemically protected form. The term “chemicallyprotected form” is used herein in the conventional chemical sense andpertains to a compound in which one or more reactive functional groupsare protected from undesirable chemical reactions under specifiedconditions (e.g., pH, temperature, radiation, solvent, and the like). Inpractice, well known chemical methods are employed to reversibly renderunreactive a functional group, which otherwise would be reactive, underspecified conditions. In a chemically protected form, one or morereactive functional groups are in the form of a protected or protectinggroup (also known as a masked or masking group or a blocked or blockinggroup). By protecting a reactive functional group, reactions involvingother unprotected reactive functional groups can be performed, withoutaffecting the protected group; the protecting group may be removed,usually in a subsequent step, without substantially affecting theremainder of the molecule. See, for example, Protective Groups inOrganic Synthesis (T. Green and P. Wuts; 3rd Edition; John Wiley andSons, 1999).

A wide variety of such “protecting,” “blocking,” or “masking” methodsare widely used and well known in organic synthesis. For example, acompound which has two nonequivalent reactive functional groups, both ofwhich would be reactive under specified conditions, may bederivatized-to render one of the functional groups “protected,” andtherefore unreactive, under the specified conditions; so protected, thecompound may be used as a reactant which has effectively only onereactive functional group. After the desired reaction (involving theother functional group) is complete, the protected group may be“deprotected” to return it to its original functionality.

For example, a hydroxy group may be protected as an ether (—OR) or anester (—OC(═O)R), for example, as: a t-butyl ether; a benzyl, benzhydryl(diphenylmethyl), or trityl (triphenylmethyl) ether; a trimethylsilyl ort-butyidimethylsilyl ether; or an acetyl ester (—OC(═O)CH₃, —OAc).

For example, an aldehyde or ketone group may be protected as an acetal(R—CH(OR)₂) or ketal (R₂C(OR)₂), respectively, in which the carbonylgroup (>C═O) is converted to a diether (>C(OR)₂), by reaction with, forexample, a primary alcohol. The aldehyde or ketone group is readilyregenerated by hydrolysis using a large excess of water in the presenceof acid.

For example, an amine group may be protected, for example, as an amide(—NRCO—R) or a urethane (—NRCO—OR), for example, as: a methyl amide(—NHCO—CH₃); a benzyloxy amide (—NHCO—OCH₂C₆H₅, —NH-Cbz); as a t-butoxyamide (—NHCO—OC(CH₃)₃, —NH-Boc); a 2-biphenyl-2-propoxy amide(—NHCO—OC(CH₃)₂C₆H₄C₆H₅, —NH-Bpoc), as a 9-fluorenylmethoxy amide(—NH-Fmoc), as a 6-nitroveratryloxy amide (—NH-Nvoc), as a2-trimethylsilylethyloxy amide (—NH-Teoc), as a 2,2,2-trichloroethyloxyamide (—NH-Troc), as an allyloxy amide (—NH-Alloc), as a2(-phenylsulphonyl)Ethyloxy amide (—NH-Psec); or, in suitable cases(e.g., cyclic amines), as a nitroxide radical (>N—O.).

For example, a carboxylic acid group may be protected as an ester forexample, as: an C₁₋₇alkyl ester (e.g., a methyl ester; a t-butyl ester);a C₁₋₇haloalkyl ester (e.g., a C₁₋₇trihaloalkyl ester); atriC₁₋₇alkylsilyl-C₁₋₇alkyl ester; or a C₅₋₂₀aryl-C₁₋₇alkyl ester (e.g.,a benzyl ester; a nitrobenzyl ester); or as an amide, for example, as amethyl amide.

For example, a thiol group may be protected as a thioether (—SR), forexample, as: a benzyl thioether; an acetamidomethyl ether(—S—CH₂NHC(═O)CH₃).

It may be convenient or desirable to prepare, purify, and/or handle theactive compound in the form of a prodrug. The term “prodrug,” as usedherein, pertains to a compound which, when metabolised (e.g., in vivo),yields the desired active compound. Typically, the prodrug is inactive,or less active than the active compound, but may provide advantageoushandling, administration, or metabolic properties.

For example, some prodrugs are esters of the active compound (e.g., aphysiologically acceptable metabolically labile ester). Duringmetabolism, the ester group (—C(═O)OR) is cleaved to yield the activedrug. Such esters may be formed by esterification, for example, of anyof the carboxylic acid groups (—C(═O)OH) in the parent compound, with,where appropriate, prior protection of any other reactive groups presentin the parent compound, followed by deprotection if required.

Examples of such metabolically labile esters include those of theformula —C(═O)OR wherein R is:

-   C₁₋₇alkyl-   (e.g., -Me, -Et, -nPr, -iPr, -nBu, -sBu, -iBu, -tBu);-   C₁₋₇aminoalkyl-   (e.g., aminoethyl; 2-(N,N-diethylamino)Ethyl;    2-(4-morpholino)Ethyl); and-   acyloxy-C₁₋₇alkyl-   (e.g., acyloxymethyl;-   acyloxyethyl;-   pivaloyloxymethyl;-   acetoxymethyl;-   1-acetoxyethyl;-   1-(1-methoxy-1-methyl)Ethyl-carbonxyloxyethyl;-   1-(benzoyloxy)Ethyl; isopropoxy-carbonyloxymethyl;-   1-isopropoxy-carbonyloxyethyl; cyclohexyl-carbonyloxymethyl;-   1-cyclohexyl-carbonyloxyethyl;-   cyclohexyloxy-carbonyloxymethyl;-   1-cyclohexyloxy-carbonyloxyethyl;-   (4-tetrahydropyranyloxy) carbonyloxymethyl;-   1-(4-tetrahydropyranyloxy)carbonyloxyethyl;-   (4-tetrahydropyranyl)carbonyloxymethyl; and-   1-(4-tetrahydropyranyl)carbonyloxyethyl).

Also, some prodrugs are activated enzymatically to yield the activecompound, or a compound which, upon further chemical reaction, yieldsthe active compound (for example, as in ADEPT, GDEPT, LIDEPT, etc.). Forexample, the prodrug may be a sugar derivative or other glycosideconjugate, or may be an amino acid ester derivative.

Acronyms

For convenience, many chemical moieties are represented using well knownabbreviations, including but not limited to, methyl (Me), ethyl (Et),n-propyl (nPr), iso-propyl (iPr), n-butyl (nBu), sec-butyl (sBu),iso-butyl (iBu), tert-butyl (tBu), n-hexyl (nHex), cyclohexyl (cHex),phenyl (Ph), biphenyl (biPh), benzyl (Bn), naphthyl (naph), methoxy(MeO), ethoxy (EtO), benzoyl (Bz), and acetyl (Ac).

For convenience, many chemical compounds are represented using wellknown abbreviations, including but not limited to, methanol (MeOH),ethanol (EtOH), iso-propanol (i-PrOH), methyl ethyl ketone (MEK), etheror diethyl ether (Et₂O), acetic acid (AcOH), dichloromethane (methylenechloride, DCM), acetonitrile (ACN), trifluoroacetic acid (TFA),dimethylformamide (DMF), tetrahydrofuran (THF), and dimethylsulfoxide(DMSO).

Synthesis of Alkane Diols Derivatives

Compounds suitable for use in the present invention may be synthesisedusing known methods. Suitable reagents and intermediates arecommercially available. Additional compounds. Additionally, severalmethods for the chemical synthesis of suitable compounds (e.g., alkanediol esters) for use in present invention are described herein. Thesemethods may be modified and/or adapted in known ways in order tofacilitate the synthesis of additional compounds suitable for use in thepresent invention.

Examples of some suitable methods for the synthesis of alkane diolmonoesters are described below.

In one method, esters of alkane diols are prepared by the reaction ofthe alkane diol with acyl halide (e.g., acyl chloride), optionally inthe presence of a base (e.g., pyridine). For example, the alkane diolmay first be dissolved in pyridine, and then the acyl halide added. Ifan excess of alkane diol is used, the mono-protected product ispredominant; if an excess of acyl halide is used, the di-protectedproduct is predominant. An example of such a method is illustrated inthe following scheme.

In another method, esters of alkane diols are prepared by the reactionof the alkane diol with a carboxylic acid, in the presence of a strongacid (e.g., H₂SO₄). For example, a small (catalytic) amount of strongacid may be used. If an excess of alkane diol is used, themono-protected product is predominant; if an excess of carboxylic acidis used, the di-protected product is predominant. An example of such amethod is illustrated in the following scheme.

In another method, esters of alkane diols are prepared by the reactionof the alkane diol with an excess of acyl anhydride, in the presence ofa base (e.g., pyridine). If an excess of alkane diol is used, themono-protected product is predominant; if an excess of acyl anhydride isused, the di-protected product is predominant. An example of such amethod is illustrated in the following scheme.

In another method, mixed esters of alkane diols are prepared by reactionof a mono-ester with an excess of acyl anhydride, in the presence of abase (e.g., pyridine). An example of such a method is illustrated in thefollowing scheme.

Suitable carboxylic acids may be prepared, for example, by reaction withaluminium trichloride (AlCl₃) and acetyl chloride (CH₃COCl) to give thecorresponding methyl ketone, which is then reacted with NaOBr (formed byreaction of Br₂ with NaOH) to give the corresponding carboxylic acid. Anexample of such a method is illustrated in the following scheme.

Suitable carboxylic acids may also be prepared by forming a Grignardreagent, which is then reacted with a borate, e.g., B(OMe)₃ to form aborane, which is then reacted with a suitable halide compound, in thepresence of a suitable catalyst, e.g, PdCl₂, to yield the desiredcarboxylic acid. An example of such a method is illustrated in thefollowing scheme.

Suitable acyl halides may be prepared, for example, by reaction of thecorresponding carboxylic acid with sulfonyl halide, e.g., sulfonylchloride (SOCl₂). An example of such a method is illustrated in thefollowing scheme.

1,4-Butanediol mono(fluorobiphenyl-4-carboxylic acid)Esters can besynthesized by an analogous method, using a commercially availableboronic acid (e.g., a fluorinated phenylboronic acid) and an iodobenzoicacid.

Commercially available fluorinated phenylboronic acids include, but arenot limited to, 2,3-difluoro-; 2,4-difluoro-; 2,5-difluoro-;2,6-difluoro-; 3,4-difluoro-; 3,5-difluoro-; 2,3,6-trifluoro-; and2,4,6-trifluoro-phenylboronic acid (Sigma-Aldrich); as well as2-fluoro-4-iodo-; 4-fluoro-3-methyl-; and 3,5-dibromo-phenylboronic acid(Lancaster). An example of such a method is illustrated in the followingscheme. In one method, iodobenzoic acid (1.75 mmol),3,4-difluorophenylboronic acid (3.5 mmol) and K₂CO₃ (2.6 mmol) arestirred in toluene (17 ml). Pd(PPh₃)₄ (0.05 mmol) is added and themixture stirred for 2 hours at 85° C. After cooling, the mixture isdiluted with ethyl acetate (17 ml), washed with saturated Na₂CO₃ (20ml), water (20 ml), 10% citric acid (20 ml), water (20 ml) and saturatedNaCl (20 ml). The solvent is evaporated and the product is purified bycolumn chromatography. In another method, a suspension of Pd(PPh₃)₄(0.05 mmol) in dimethoxyethanol (20 ml) is prepared. Iodobenzoic acid (2mmol) is added and the mixture is stirred for 10 minutes.3,4-Difluorophenylboronic acid (3 mmol) in ethanol (2 ml) is addedfollowed by 2 M Na₂CO₃ (4 mmol). The mixture is refluxed for 18 hours,filtered and evaporated. The residue is washed with saturated NaCl (20ml) and product is purified by column chromatography. See, for example,Miyaura et al., 1995.

Methods for the preparation of compounds having a phosphonic acid group,include, for example, those described below.

In one method, ethenylidenebisphosphonate (CH₂═C(P(═O)(OR)₂)₂) isprepared from paraformaldehyde, diethylamine and a tetraalkyl methylenebisphosphonate (H₂C(P(═O)(OR)₂)₂), using, for example, the methoddescribed by Degenhardt and Burdsall, 1986. Theethenylidene-bisphosphonate is then reacted with, for example, ABD-0056(4BP), in methylene chloride, in the presence of triethylamine, using,for example, the method described by Herczegh et al., 2002. Thephosphate ester groups, e.g., ethyl groups, are removed e.g., withtrimethylsilylbromide or left in place. An example of such a method isillustrated in the following scheme.

In another method, ABD-0056, for example, is heated withtriethylorthoformate and diethyl phosphite (HP(═O)(OEt)₂) using, forexample, the method described by Herczegh et al., 2002. Again, thephosphate ester groups, e.g., ethyl groups, are removed e.g., withtrimethylsilylbromide or left in place. An example of such a method isillustrated in the following scheme.

In another method, 4-bromobutanol (prepared, for example, by Method 9described herein) is acetylated in acetic anhydride/pyridine. Theresultant 4-acetoxybutylbromide is then heated with triethylphosphite togive diethyl-4-acetoxybutyl phosphonate using, for example, the methoddescribed by Eberhard and Westheimer, 1965. Hydrolysis with sulphuricacid removes the acetyl and ethyl groups to give the4-hydroxybutylphosphonate. This is then linked to biphenyl-4-carboxylicacid using N-methyl morpholine and isobutyl chloroformate in a mixtureof tetrahydrafuran and dimethylformamide. An example of such a method isillustrated in the following scheme.

In another method, 1,4-dibromobutane is heated with triethylphosphite togive diethyl-4-bromobutylphosphonate, for example, as described byEberhard and Westheimer, 1965. The resultant bromide is then reactedwith, for example, biphenyl-4-carboxylic acid in dimethylformamide, inthe presence of potassium carbonate. Again, the phosphate ester groups,e.g., ethyl groups, are removed e.g., with trimethylsilylbromide or leftin place. An example of such a method is illustrated in the followingscheme.

In another method, ABD-0086 (4BP-Br), for example, is gently refluxedwith triisopropylphosphite to give the phosphonylated product. Remainingtriisopropylphosphite is removed by distillation under reduced pressureand the residue is purified by column chromatography to give a clearoil. The isopropyl groups are removed using trimethylsilylbromide indichloromethane, or left in place. An example of such a method isillustrated in the following scheme.

In another method, acrylic acid methyl ester (methyl acrylate) andmethylene diphosphonic acid tetraethyl ester are mixed and saturatedsodium ethanolate solution is added dropwise. The mixture is heated(e.g., to 90° C. for 2 hours) and the product obtained by distillationunder reduced pressure. The ester groups were removed by hydrolysis inconc. HCl. See, e.g., Blum et al., 1978. An example of such a method isillustrated in the following scheme.

The products may be purified, for example, by column chromatography.

Use of Alkane Diol Derivatives

The present invention provides active compounds, specifically, activealkane diol derivatives (e.g., esters of alkane diols), as describedherein, which inhibit osteoclasts, for example, inhibit of the survival,formation, and/or activity of osteoclasts, and/or which inhibit boneresorption. The compounds may therefore be referred to as “osteoclastinhibitors” and/or “bone resorption inhibitors.”

The term “active,” as used herein, pertains to compounds which arecapable of inhibiting the survival, formation, and/or activity ofosteociasts, and/or inhibiting bone resorption, and specificallyincludes both compounds with intrinsic activity (drugs) as well asprodrugs of such compounds, which prodrugs may themselves exhibit littleor no intrinsic activity.

One of ordinary skill in the art is readily able to determine whether ornot a candidate compound inhibits the survival, formation, and/oractivity of osteoclasts and/or inhibits bone resorption. For example,suitable methods which may conveniently be used in order to assess theinhibitory effects offered by a particular compound are described in theexamples below.

The compounds of the present invention are also useful in the treatmentof conditions mediated by osteoclasts (as “osteoclast inhibitors”),and/or conditions characterised by bone resorption (as “bone resorptioninhibitors”). Examples of such conditions include, but are not limitedto, the following:

Diseases of the skeleton, including but not limited to, pathologicallylow bone mineral density, such as osteoporosis (including, e.g., steroidinduced osteoporosis); osteopetrosis; osteoarthritis; ectopic boneformation; Paget's disease of bone (osteitis deformans); and rheumatoidarthritis.

Neoplasia of bones, both as a primary tumour and as metastases,including but not limited to, osteosarcoma and osteoma (Zheng et al.,1998, J. Cell Biochem., Vol. 70, p. 121) and cancer associated bonedisease (e.g., multiple myeloma).

Examples of preferred conditions include osteoporosis, rheumatoidarthritis, cancer associated bone disease, and Paget's disease.

The compounds of the present invention have also macrophage inhibitoryeffects, and so are useful in the treatment of conditions associatedwith inflammation or activation of the immune system. Examples of suchconditions include, but are not limited to, the following: Diseases withan inflammatory or autoimmune component, including allergic diseases,such as atopy, allergic rhinitis, atopic dermatitis, anaphylaxis,allergic bronchopulmonary aspergillosis, and hypersensitivitypneumonitis (pigeon breeders disease, farmer's lung disease, humidifierlung disease, malt workers' lung disease); allergies, including fleaallergy dermatitis in mammals such as domestic animals, e.g., dogs andcats, contact allergens including mosquito bites or other insect stingallergies, poison ivy, poison oak, poison sumac, or other skinallergens; autoimmune disorders, including, but not limited to, type Idiabetes, Crohn's disease, multiple sclerosis, arthritis, rheumatoidarthritis (Ogata et at., 1997, J. Pathol., Vol. 182, p. 106); Gong etal., 1997, J. Exp. Med., Vol 186, p. 131), systemic lupus erythematosus,autoimmune (Hasimoto's) thyroiditis, autoimmune liver diseases such ashepatitis and primary biliary cirrhosis, hyperthyroidism (Graves'disease; thyrotoxicosis), insulin-resistant diabetes, autoimmune adrenalinsufficiency (Addison's disease), autoimmune oophoritis, autoimmuneorchitis, autoimmune hemolytic anemia, paroxysmal cold hemoglobinuria,Behcet's disease, autoimmune thrombocytopenia, autoimmune neutropenia,pernicious anemia, pure red cell anemia, autoimmune coagulopathies,myasthenia gravis, experimental allergic encephalomyelitis, autoimmunepolyneuritis, pemphigus and other bullous diseases, rheumatic carditis,Goodpasture's syndrome, postcardiotomy syndrome, Sjogren's syndrome,polymyositis, dermatomyositis, and scleroderma; disease states resultingfrom inappropriate inflammation, either local or systemic, for example,irritable or inflammatory bowel syndrome (Mazzucchelli et al., 1996, J.Pathol., Vol. 178, p. 201), skin diseases such as psoriasis and lichenplanus, delayed type hypersensitivity, chronic pulmonary inflammation,e.g., pulmonary alveolitis and pulmonary granuloma, gingivalinflammation or other periodontal disease, and osseous inflammationassociated with lesions of endodontic origin (Volejnikova et al., 1997,Am. J. Pathol., Vol. 150, p. 1711), hypersensitivity lung diseases suchas hypersensitivity pneumonitis (Sugiyama et al., 1995, Eur. Respir. J.,Vol. 8, p. 1084), and inflammation related to histamine release frombasophils (Dvorak et al., 1996, J. Allergy Clin. Immunol., Vol. 98, p.355), such as hay fever, histamine release from mast cells (Galli etal., 1989, Ciba Foundation Symposium, Vol. 147, p. 53), or mast celltumors, types of type 1 hypersensitivity reactions (anaphylaxis, skinallergy, hives, allergic rhinitis, and allergic gastroenteritis);ulcerative colitis.

Thus, one aspect of the invention pertains to a method of inhibitingosteociast survival, formation, and activity, in vitro or in vivo,comprising contacting an osteoclast with an effective amount of anactive compound, as described herein.

Another aspect of the invention pertains to a method of inhibiting boneresorption, in vitro or in vivo, comprising contacting cells in the bonemicroenvironment with a therapeutically-effective amount of an activecompound, as described herein.

The term “cells in the bone microenvironment,” as used herein, pertainsto cells such as osteoblasts, osteoclasts, osteocytes and bone marrowstromal cells, which are located in close proximity to bone (e.g.,within one hundred micrometers of the bone surface).

Another aspect of the present invention pertains to a method for thetreatment of a condition mediated by osteociasts and/or characterised bybone resorption, as described herein, comprising administering to asubject suffering from said condition a therapeutically-effective amountof an active compound, as described herein, preferably in the form of apharmaceutical composition.

Another aspect of the present invention pertains to a method for thetreatment of a condition associated with inflammation or activation ofthe immune system, as described herein, comprising administering to asubject suffering from said condition a therapeutically-effective amountof an active compound, as described herein, preferably in the form of apharmaceutical composition.

Another aspect of the present invention pertains to an active compoundas described herein for use in a method of treatment of the human oranimal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionmediated by osteoclasts and/or characterised by bone resorption, asdescribed herein, of the human or animal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionmediated by osteoclasts, as described herein, of the human or animalbody by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditioncharacterised by bone resorption, as described herein, of the human oranimal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment of osteoporosis,rheumatoid arthritis, cancer associated bone disease, or Paget's diseaseof the human or animal body by therapy.

Another aspect of the present invention pertains to an active compound,as described herein, for use in a method of treatment a conditionassociated with inflammation or activation of the immune system, asdescribed herein, of the human or animal body by therapy.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition mediated by osteoclasts and/orcharacterised by bone resorption, as described herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition mediated by osteoclasts, asdescribed herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition characterised by bone resorption, asdescribed herein.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of osteoporosis, rheumatoid arthritis, cancerassociated bone disease, or Paget's disease.

Another aspect of the present invention pertains to use of an activecompound, as described herein, for the manufacture of a medicament foruse in the treatment of a condition associated with inflammation oractivation of the immune system, as described herein.

The term “treatment,” as used herein in the context of treating acondition, pertains generally to treatment and therapy, whether of ahuman or an animal (e.g., in veterinary applications), in which somedesired therapeutic effect is achieved, for example, the inhibition ofthe progress of the condition, and includes a reduction in the rate ofprogress, a halt in the rate of progress, amelioration of the condition,and cure of the condition. Treatment as a prophylactic measure (i.e.,prophylaxis) is also included.

The term “therapeutically-effective amount,” as used herein, pertains tothat amount of an active compound, or a material, composition or dosagefrom comprising an active compound, which is effective for producingsome desired therapeutic effect, commensurate with a reasonablebenefit/risk ratio, when administered in accordance with a desiredtreatment regimen.

The term “treatment” includes combination treatments and therapies, inwhich two or more treatments or therapies are combined, for example,sequentially or simultaneously. Examples of treatments and therapiesinclude, but are not limited to, chemotherapy (the administration ofactive agents, including, e.g., drugs, antibodies (e.g., as inimmunotherapy), prodrugs (e.g., as in photodynamic therapy, GDEPT,ADEPT, etc.); surgery; radiation therapy; and gene therapy.

Active compounds may also be used as cell culture additives to inhibitosteoclasts, for example, to inhibit the survival, formation, and/oractivity of osteoclasts.

Active compounds may also be used as part of an in vitro assay, forexample, in order to determine whether a candidate host is likely tobenefit from treatment with the compound in question.

Active compounds may also be used as a standard, for example, in anassay, in order to identify other active compounds, other osteoclastinhibitors, etc.

Kits

One aspect of the invention pertains to a kit comprising (a) an activecompound as described herein, or a composition comprising an activecompound as described herein, e.g., preferably provided in a suitablecontainer and/or with suitable packaging; and (b) instructions for use,e.g., written instructions on how to administer the active compound orcomposition.

The written instructions may also include a list of indications forwhich the active ingredient is a suitable treatment.

Routes of Administration

The active compound or pharmaceutical composition comprising the activecompound may be administered to a subject by any convenient route ofadministration, whether systemically/peripherally or topically (i.e., atthe site of desired action).

Routes of administration include, but are not limited to, oral (e.g., byingestion); buccal; sublingual; transdermal (including, e.g., by apatch, plaster, etc.); transmucosal (including, e.g., by a patch,plaster, etc.); intranasal (e.g., by nasal spray); ocular (e.g., byeyedrops); pulmonary (e.g., by inhalation or insufflation therapy using,e.g., via an aerosol, e.g., through the mouth or nose); rectal (e.g., bysuppository or enema); vaginal (e.g., by pessary); parenteral, forexample, by injection, including subcutaneous, intradermal,intramuscular, intravenous, intraarterial, intracardiac, intrathecal,intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal,intratracheal, subcuticular, intraarticular, subarachnoid, andintrasternal; by implant of a depot or reservoir, for example,subcutaneously or intramuscularly.

The Subject

The subject may be a chordate, a vertebrate, a mammal, a bird, a reptile(e.g., snakes, lizards, crocodiles), an amphibian (e.g., frogs, toads),a bony fish (e.g., salmon, plaice, eel, lungfish), a cartilaginous fish(e.g., sharks, rays), or a jawless fish (e.g., lampreys, hagfish).

The subject may be a mammal, a placental mammal, a marsupial (e.g.,kangaroo, wombat), a monotreme (e.g., duckbilled platypus), a rodent(e.g., a guinea pig, a hamster, a rat, a mouse), murine (e.g., a mouse),a lagomorph (e.g., a rabbit), avian (e.g., a bird), canine (e.g., adog), feline (e.g., a cat), equine (e.g., a horse), porcine (e.g., apig), ovine (e.g., a sheep), bovine (e.g., a cow), a primate, simian(e.g., a monkey or ape), a monkey (e.g., marmoset, baboon), an ape(e.g., gorilla, chimpanzee, orangutang, gibbon), or a human.

Furthermore, the subject may be any of its forms of development, forexample, a foetus.

In one preferred embodiment, the subject is a human.

Formulations

While it is possible for the active compound to be administered alone,it is preferable to present it as a pharmaceutical formulation (e.g.,composition, preparation, medicament) comprising at least one activecompound, as defined above, together with one or more otherpharmaceutically acceptable ingredients well known to those skilled inthe art, including, but not limited to, pharmaceutically acceptablecarriers, diluents, excipients, adjuvants, fillers, buffers,preservatives, anti-oxidants, lubricants, stabilisers, solubilisers,surfactants (e.g., wetting agents), masking agents, colouring agents,flavouring agents, and sweetening agents. The formulation may furthercomprise other active agents, for example, other therapeutic orprophylactic agents.

Thus, the present invention further provides pharmaceuticalcompositions, as defined above, and methods of making a pharmaceuticalcomposition comprising admixing at least one active compound, as definedabove, together with one or more other pharmaceutically acceptableingredients well known to those skilled in the art, e.g., carriers,diluents, excipients, etc. If formulated as discrete units (e.g.,tablets, etc.), each unit contains a predetermined amount (dosage) ofthe active compound.

The term “pharmaceutically acceptable” as used herein pertains tocompounds, ingredients, materials, compositions, dosage forms, etc.,which are, within the scope of sound medical judgment, suitable for usein contact with the tissues of the subject in question (e.g., human)without excessive toxicity, irritation, allergic response, or otherproblem or complication, commensurate with a reasonable benefit/riskratio. Each carrier, diluent, excipient, etc. must also be “acceptable”in the sense of being compatible with the other ingredients of theformulation.

Suitable carriers, diluents, excipients, etc. can be found in standardpharmaceutical texts, for example, Remington's Pharmaceutical Sciences,18th edition, Mack Publishing Company, Easton, Pa., 1990; and Handbookof Pharmaceutical Excipients, 2nd edition, 1994.

The formulations may be prepared by any methods well known in the art ofpharmacy. Such methods include the step of bringing into association theactive compound with a carrier which constitutes one or more accessoryingredients. In general, the formulations are prepared by uniformly andintimately bringing into association the active compound with carriers(e.g., liquid carriers, finely divided solid carrier, etc.), and thenshaping the product, if necessary.

The formulation may be prepared to provide for rapid or slow release;immediate, delayed, timed, or sustained release; or a combinationthereof.

Formulations may suitably be in the form of liquids, solutions (e.g.,aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous),emulsions (e.g., oil-in-water, water-in-oil), elixirs, syrups,electuaries, mouthwashes, drops, tablets (including, e.g., coatedtablets), granules, powders, losenges, pastilles, capsules (including,e.g., hard and soft gelatin capsules), cachets, pills, ampoules,boluses, suppositories, pessaries, tinctures, gels, pastes, ointments,creams, lotions, oils, foams, sprays, mists, or aerosols.

Formulations may suitably be provided as a patch, adhesive plaster,bandage, dressing, or the like which is impregnated with one or moreactive compounds and optionally one or more other pharmaceuticallyacceptable ingredients, including, for example, penetration, permeation,and absorption enhancers. Formulations may also suitably be provided inthe form of a depot or reservoir.

The active compound may be dissolved in, suspended in, or admixed withone or more other pharmaceutically acceptable ingredients. The activecompound may be presented in a liposome or other microparticulate whichis designed to target the active compound, for example, to bloodcomponents or one or more organs.

Formulations suitable for oral administration (e.g, by ingestion)include liquids, solutions (e.g., aqueous, non-aqueous), suspensions(e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water,water-in-oil), elixirs, syrups, electua des, tablets, granules, powders,capsules, cachets, pills, ampoules, boluses.

Formulations suitable for buccal administration include mouthwashes,losenges, pastilles, as well as patches, adhesive plasters, depots, andreservoirs. Losenges typically comprise the active compound in aflavored basis, usually sucrose and acacia or tragacanth. Pastillestypically comprise the active compound in an inert matrix, such asgelatin and glycerin, or sucrose and acacia. Mouthwashes typicallycomprise the active compound in a suitable liquid carrier.

Formulations suitable for sublingual administration include tablets,losenges, pastilles, capsules, and pills.

Formulations suitable for oral transmucosal administration includeliquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g.,aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil),mouthwashes, losenges, pastilles, as well as patches, adhesive plasters,depots, and reservoirs.

Formulations suitable for non-oral transmucosal administration includeliquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g.,aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil),suppositories, pessaries, gels, pastes, ointments, creams, lotions,oils, as well as patches, adhesive plasters, depots, and reservoirs.

Formulations suitable for transdermal administration include gels,pastes, ointments, creams, lotions, and oils, as well as patches,adhesive plasters, bandages-, dressings, depots, and reservoirs.

Tablets may be made by conventional means, e.g., compression or molding,optionally with one or more accessory ingredients. Compressed tabletsmay be prepared by compressing in a suitable machine the active compoundin a free-flowing form such as a powder or granules, optionally mixedwith one or more binders (e.g., povidone, gelatin, acacia, sorbitol,tragacanth, hydroxypropylmethyl cellulose); fillers or diluents (e.g.,lactose, microcrystalline cellulose, calcium hydrogen phosphate);lubricants (e.g., magnesium stearate, talc, silica); disintegrants(e.g., sodium starch glycolate, cross-linked povidone, cross-linkedsodium carboxymethyl cellulose); surface-active or dispersing or wettingagents (e.g., sodium lauryl sulfate); preservatives (e.g., methylp-hydroxybenzoate, propyl p-hydroxybenzoate, sorbic acid); flavours,flavour enhancing agents, and sweeteners. Molded tablets may be made bymolding in a suitable machine a mixture of the powdered compoundmoistened with an inert liquid diluent. The tablets may optionally becoated or scored and may be formulated so as to provide slow orcontrolled release of the active compound therein using, for example,hydroxypropylmethyl cellulose in varying proportions to provide thedesired release profile. Tablets may optionally be provided with acoating, for example, to affect release, for example an enteric coating,to provide release in parts of the gut other than the stomach.

Ointments are typically prepared from the active compound and aparaffinic or a water-miscible ointment base.

Creams are typically prepared from the active compound and anoil-in-water cream base. If desired, the aqueous phase of the cream basemay include, for example, at least about 30% w/w of a polyhydricalcohol, i.e., an alcohol having two or more hydroxyl groups such aspropylene glycol, butane-1,3-diol, mannitol, sorbitol, glycerol andpolyethylene glycol and mixtures thereof. The topical formulations maydesirably include a compound which enhances absorption or penetration ofthe active compound through the skin or other affected areas. Examplesof such dermal penetration enhancers include dimethylsulfoxide andrelated analogues.

Emulsions are typically prepared from the active compound and an oilyphase, which may optionally comprise merely an emulsifier (otherwiseknown as an emulgent), or it may comprises a mixture of at least oneemulsifier with a fat or an oil or with both a fat and an oil.Preferably, a hydrophilic emulsifier is included together with alipophilic emulsifier which acts as a stabiliser. It is also preferredto include both an oil and a fat. Together, the emulsifier(s) with orwithout stabiliser(s) make up the so-called emulsifying wax, and the waxtogether with the oil and/or fat make up the so-called emulsifyingointment base which forms the oily dispersed phase of the creamformulations.

Suitable emulgents and emulsion stabilisers include Tween 60, Span 80,cetostearyl alcohol, myristyl alcohol, glyceryl monostearate and sodiumlauryl sulphate. The choice of suitable oils or fats for the formulationis based on achieving the desired cosmetic properties, since thesolubility of the active compound in most oils likely to be used inpharmaceutical emulsion formulations may be very low. Thus the creamshould preferably be a non-greasy, non-staining and washable productwith suitable consistency to avoid leakage from tubes or othercontainers. Straight or branched chain, mono- or dibasic alkyl esterssuch as di-isoadipate, isocetyl stearate, propylene glycol diester ofcoconut fatty acids, isopropyl myristate, decyl oleate, isopropylpalmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branchedchain esters known as Crodamol CAP may be used, the last three beingpreferred esters. These may be used alone or in combination depending onthe properties required. Alternatively, high melting point lipids suchas white soft paraffin and/or liquid paraffin or other mineral oils canbe used.

Formulations suitable for intranasal administration, where the carrieris a liquid, include, for example, nasal spray, nasal drops, or byaerosol administration by nebuliser, include aqueous or oily solutionsof the active compound.

Formulations suitable for intranasal administration, where the carrieris a solid, include, for example, those presented as a coarse powderhaving a particle size, for example, in the range of about 20 to about500 microns which is administered in the manner in which snuff is taken,i.e., by rapid inhalation through the nasal passage from a container ofthe powder held close up to the nose.

Formulations suitable for pulmonary administration (e.g., by inhalationor insufflation therapy) include those presented as an aerosol sprayfrom a pressurised pack, with the use of a suitable propellant, such asdichlorodifluoromethane, trichlorofluoromethane,dichoro-tetrafluoroethane, carbon dioxide, or other suitable gases.

Formulations suitable for ocular administration include eye dropswherein the active compound is dissolved or suspended in a suitablecarrier, especially an aqueous solvent for the active compound.

Formulations suitable for rectal administration may be presented as asuppository with a suitable base comprising, for example, natural orhardened oils, waxes, fats, semi-liquid or liquid polyols, for example,cocoa butter or a salicylate; or as a solution or suspension fortreatment by enema.

Formulations suitable for vaginal administration may be presented aspessaries, tampons, creams, gels, pastes, foams or spray formulationscontaining in addition to the active compound, such carriers as areknown in the art to be appropriate.

Formulations suitable for parenteral administration (e.g., byinjection), include aqueous or non-aqueous, isotonic, pyrogen-free,sterile liquids (e.g., solutions, suspensions), in which the activecompound is dissolved, suspended, or otherwise provided (e.g., in aliposome or other microparticulate). Such liquids may additional containother pharmaceutically acceptable ingredients, such as anti-oxidants,butters, preservatives, stabilisers, bacteriostats, suspending agents,thickening agents, and solutes which render the formulation isotonicwith the blood (or other relevant bodily fluid) of the intendedrecipient. Examples of excipients include, for example, water, alcohols,polyols, glycerol, vegetable oils, and the like. Examples of suitableisotonic carriers for use in such formulations include Sodium ChlorideInjection, Ringer's Solution, or Lactated Ringer's Injection. Typically,the concentration of the active compound in the liquid is from about 1ng/ml to about 10 μg/ml, for example from about 10 ng/ml to about 1μg/ml. The formulations may be presented in unit-dose or multi-dosesealed containers, for example, ampoules and vials, and may be stored ina freeze-dried (lyophilised) condition requiring only the addition ofthe sterile liquid carrier, for example water for injections,immediately prior to use. Extemporaneous injection solutions andsuspensions may be prepared from sterile powders, granules, and tablets.

Dosage

It will be appreciated by one of skill in the art that appropriatedosages of the active compounds, and compositions comprising the activecompounds, can vary from patient to patient. Determining the optimaldosage will generally involve the balancing of the level of therapeuticbenefit against any risk or deleterious side effects. The selecteddosage level will depend on a variety of factors including, but notlimited to, the activity of the particular compound, the route ofadministration, the time of administration, the rate of excretion of thecompound, the duration of the treatment, other drugs, compounds, and/ormaterials used in combination, the severity of the condition, and thespecies, sex, age, weight, condition, general health, and prior medicalhistory of the patient. The amount of compound and route ofadministration will ultimately be at the discretion of the physician,veterinarian, or clinician, although generally the dosage will beselected to achieve local concentrations at the site of action whichachieve the desired effect without causing substantial harmful ordeleterious side-effects.

Administration can be effected in one dose, continuously orintermittently (e.g., in divided doses at appropriate intervals)throughout the course of treatment. Methods of determining the mosteffective means and dosage of administration are well known to those ofskill in the art and will vary with the formulation used for therapy,the purpose of the therapy, the target cell(s) being treated, and thesubject being treated. Single or multiple administrations can be carriedout with the dose level and pattern being selected by the treatingphysician, veterinarian, or clinician.

In general, a suitable dose of the active compound is in the range ofabout 100 pg to about 250 mg (more typically about 100 pg to about 25mg) per kilogram body weight of the subject per day. Where the activecompound is a salt, an ester, an amide, a prodrug, or the like, theamount administered is calculated on the basis of the parent compoundand so the actual weight to be used is increased proportionately.

EXAMPLES

The following are examples are provided solely to illustrate the presentinvention and are not intended to limit the scope of the invention, asdescribed herein.

The identity and purity of the compounds was proven using ¹³C and ¹H nmrin comparison with model literature compounds (ABD-0006 and ABD-0009)(4A and 4B), for which accurate melting or boiling points can also beobtained. The identity of many compounds was also ascertained using gaschromatography-mass spectroscopy (GC-MS).

General Methods

Method 1: Esterification Using an Acid Chloride and an Alcohol

The alcohol (0.1 mol) was dissolved in dry pyridine (50 ml) and chilledin an ice bath. The acid chloride (0.02 mol) was added dropwise withvigorous stirring and the mixture stirred overnight at room temperature.The mixture was poured into water (200 ml) and extracted with methylenechloride (100 ml). The organic phase was washed sequentially with 2 MHCl (3×100 ml), water, saturated NaHCO₃ and water. The solution wasdried over Na₂SO₄ and evaporated to an oil or amorphous solid. Theproduct was dissolved in methylene chloride (<5 ml), absorbed onto acolumn (silica gel 60, Merck) and purified using a mixture of lightpetroleum and ethyl acetate (usually a 1:1 mixture was satisfactory).Evaporation of the solvent gave an oil or a solid.

Method 2: Esterification Using an Acid and an Alcohol

The alcohol (0.2 mol) was heated in a boiling water bath. The acid (0.02mol) was added followed by 20 drops of concentrated H₂SO₄ with vigorousstirring. Stirring was continued for 3 hours or until all of the acidhad dissolved. The solution was poured into water (200 ml) and extractedwith methylene chloride. The organic phase was washed with water,saturated NaHCO₃ and water. The solution was dried over Na₂SO₄ andpurified as described in Method 1.

Method 3: Esterification Using an Acid Anhydride and an Alcohol

Pyridine (25 ml) and acetic anhydride (25 ml) were stirred at 0° C. Thealcohol (4 mmol) in pyridine (10 ml) was added dropwise. The mixture wasstirred overnight at room temperature and then in a boiling water bathfor 2 hours. The mixture was then poured into water (200 ml) andextracted with methylene chloride (100 ml). The organic phase was washedsequentially with water, 2 M HCl (2×100 ml), water, saturated NaHCO₃(100 ml) and water. The organic phase was dried over Na₂SO₄, evaporatedand purified by column chromatography (light petroleum:ethyl acetate,3:1).

Method 4: Preparation of Biphenylcarboxylates by Friedel-CraftsAcylation

The biphenyl (0.03 mol) was added to 1 M AlCl₃ in nitrobenzene (40 ml,0.04 mol) with chilling in an ice bath. Acetyl chloride (0.06 mol) wasadded dropwise and the mixture stirred overnight at room temperature.The dark solution was poured into a mixture of crushed ice (150 ml),water (25 ml) and conc. HCl (50 ml). The organic phase was separated andthe nitrobenzene removed by steam distillation to give a dark lowmelting solid. The solid was recrystallised from aqueous methanol.

Method 5: Oxidation of Acetate Group to Free Acid

NaOH (7 g) was dissolved in water (25 ml) and cooled in an ice bath.Bromine (7.8 g) was added dropwise to give a solution of NaOBr. Thebiphenyl acetate from Method 4 (0.01 mol) was dissolved in dioxane (35ml) and warmed to 50° C. in a water bath. The NaOBr solution was addedslowly to the stirred solution of the biphenyl acetate and stirringcontinued at 50° C. for a further 20 minutes. The solution was allowedto cool and a solution of sodium metabisulphite (Na₂S₂O₅) (8 g in 40 mlwater) was added followed by water (170 ml). 50 ml of the liquid wasevaporated under reduced pressure with heating. The remainder wasacidified with conc._HCl (5 ml) and a white precipitate formed uponcooling. The precipitate was filtered and recrystallised from aceticacid.

Method 6: Preparation of Biphenyls by Suzuki Coupling

The substituted benzylbromide (e.g., 4-bromotoluene) (16 mmol) wasdissolved in dry ether (15 ml) and reacted with magnesium (0.4 g, 16mmol) to form the Grignard reagent. Gentle heating may be required toinitiate the reaction. Trimethylborate (0.42 g, 4 mmol) was dissolved inether (5 ml). The Grignard reagent was added dropwise to this solutionwith vigorous stirring. The reaction mixture was boiled for 15 minutesto give the aryl borane. A solution containing NaOH (2 g), 4-iodobenzoicacid (10 mmol), and PdCl₂ (0.1 mmol) in water (70 ml) was prepared andadded dropwise to the aryl borane with vigorous stirring. Followingaddition, the mixture was boiled for 1 hour, allowed to cool andextracted with ether. The aqueous was extracted with methylene chloride.The aqueous was then acidified with HCl and extracted with methylenechloride followed by diethyl ether. These final two fractions werewashed with saturated NaHCO₃ solution and water. All four fractions werecombined, dried over Na₂SO₄ and evaporated to an amorphous solid.Purification by column chromatography (ethyl acetate:light petroleum,1:1) gave the desired product as a white powder.

Method 7: Acetylation

The alcohol (10 mmol) was dissolved in pyridine (25 ml) and aceticanhydride (10 ml) was added dropwise. The mixture was stirred overnight,poured into water (200 ml) and extracted with methylene chloride (100ml). The organic phase was washed sequentially with water, 2M HCl (2×100ml), water, saturated NaHCO₃ (100 ml) and water. The organic phase wasdried over Na₂SO₄, evaporated and purified by column chromatography(light petroleum:ethyl acetate, 3:1).

Method 8: Preparation of Acid Chloride

The acid (10 mmol) was dissolved in thionyl chloride (SOCl₂) (30 ml) andrefluxed for 3 hours. The mixture was poured into acetic acid (100 ml)and left to stand until bubbling ceased. Volatile components wereremoved under vacuum and the mixture left to crystallise overnight. Thedesired acid chloride was collected by filtration.

Method 9: Preparation of 4-bromobutanol

48% Hydrobromic acid (200 ml) was added to refluxing tetrahydrafuran(400 ml) over a period of 1 hour. Reflux was continued for 4 hours. Thesolution was allowed to cool, excess HBr was neutralised with NaHCO₃ andpartitioned with water. The organic phase was separated, washed withbrine and dried over MgSO₄. Evaporation of the solvent gave4-bromobutanol as an oil.

Example 1 1,4-Butanediol di(acetic acid)Ester (ABD-0006) (4A)

The title compound was prepared from acetic anhydride and 1,4-butanediolusing Method 3, and was purified by distillation under reduced pressureto give a clear oil (yield 60%). δ_(C) (CDCl₃, 62.9 MHz): 20.96, 25.3,63.9 and 171.1. δ_(H) (CDCl₃, 250 MHz): 1.70 (4H, m), 2.02 (6H, m) and4.10 (4H, m).

Example 2 1,4-Butanediol di(butanoic acid)Ester (ABD-0007) (4BU)

The title compound was prepared from butyric acid and 1,4-butanediolusing Method 2, to give a clear oil (yield 85%). δ_(C) (CDCl₃, 62.9MHz): 13.6, 18.4, 25.4, 36.2, 63.7 and 173.7. δ_(H) (CDCl₃, 250 MHz):0.92 (6H, t, J 7.3), 1.58-1.70 (8H, m), 2.26 (4H, t, J 7.3) and 4.07(4H, m).

Example 3 1,4-Butanediol di(cyclclohexanecarboxylic acid)Ester(ABD-0019) (4C)

The title compound was prepared from cyclohexanecarbonyl chloride and1,4-butanediol using Method 1 to give a clear oil which solidified overtime (yield 30%). δ_(C) (CDCl₃, 62.9 MHz): 25.5, 25.8, 29.0, 43.2, 63.6and 176.1.

Example 4 1,4-Butanediol di(benzoic acid)Ester (ABD-0009) (4B)

The title compound was prepared from benzoic acid and 1,4-butanediolusing Method 2, to give a white crystalline solid (yield 80%). δ_(C)(CDCl₃, 62.9 MHz): 25.6, 64.5, 128.4, 129.6, 130.3, 133.0 and 166.6.δ_(H) (CDCl₃, 250 MHz): 1.95 (4H, m), 4.40 (4H, m), 7.43 (4H, t, J 7.3),7.54 (2H, d, J 7.0) and 8.04 (4H, d, J 7.0).

Example 5 1,4-Butanediol di(phenylacetic acid)Ester (ABD-0014) (4P)

The title compound was prepared from phenylacetic acid and1,4-butanediol using Method 2, to give a white solid (yield 50%). δ_(C)(CDCl₃, 62.9 MHz): 19.8, 23.8, 36.3, 57.1, 59.5, 121.9, 123.4, 124.1,128.9, 157.4 and 166.5. δ_(H) (CDCl₃, 250 MHz): 1.64 (4H, m), 3.61 (4H,s), 4.08 (4H, m) and 7.29 (10H, m).

Example 6 1,6-Hexanediol di(phenylacetic acid)Ester (ABD-0017) (6P)

The title compound was prepared from phenylacetic acid and1,6-hexanediol using Method 2 to give a clear oil (yield 90%). δ_(c)(CDCl₃, 62.9 MHz): 25.5, 28.4, 41.5, 64.7, 127.1, 128.6, 129.3, 134.2,and 171.6.

Example 7 1,4-Butanediol di(pentafluorobenzoic acid)Ester (ABD-0085)(10F)

The title compound was prepared from pentafluorobenzoyl chloride and1,4-butanediol using Method 1 to give a white solid. Columnchromatography gave the title compound as the first fraction (yield20%). δ_(C) (CDCl₃, 62.9 MHz): 25.0 and 66.0.

Example 8 1,4-Butanediol di(2,4-difluorobenzoic acid)Ester (ABD-0111)(D2,4FB)

The title compound was prepared from 2,4-difluorobenzoyl chloride and1,4-butanediol using Method 1, to give a clear oil. Columnchromatography gave the title compound as the first fraction (yield20%). δ_(C) (CDCl₃, 62.9 MHz): 25.4, 64.9, 105.2 (t, J 26.4), 111.6 (dd,J 21.5, 2.9) and 133.9 (d, J 10.7).

Example 9 2,2,3,3-tetrafluorobutan-1,4-diol di(biphenyl-4-carboxylicacid)Ester (ABD-0096) (DBP-4F)

The title compound was prepared from 2,2,3,3-tetrafluorobutane-1,4-dioland biphenylcarbonyl chloride using Method 1 to give a white (yield20%). Purification by column chromatography (ethyl acetate:lightpetroleum, 1:1) gave the title compound as the first fraction. δ_(C)(CDCl₃, 62.9 MHz): 127.3, 127.3, 128.4, 129.0, 130.6, 139.8, 146.5 and165.4.

Example 10 Biphenyl-4-carboxylic acid (4-acetoxy)butyl ester (ABD-0049)(4BP-acetate)

The title compound was prepared from ABD-0056 using Method 7 to give awhite solid (yield 90%). δ_(C) (CDCl₃, 62.9 MHz): 21.0, 25.4, 25.5,64.0, 64.5, 127.1, 127.3, 128.2, 129.0, 130.1, 140.0, 145.7, 166.5 and171.2. δ_(H) (CDCl₃, 250 MHz): 1.8 (4H, s, 2×CH₂), 2.0 (3H, s, COCH₃),4.1 (2H, t, J 6.3), 4.4 (2H, t, J 6.3), 7.4 (3H, m), 7.6 (4H, 2×d) and8.1 (2H, d, J 8.8).

Example 11 1,4-Butanediol mono(benzoic acid)Ester (ABD-0008) (4 MB)

The title compound was prepared from benzoyl chloride and 1,4-butanediolusing Method 1 to give a pale oil (yield 40%). δ_(C) (CDCl₃, 62.9 MHz):25.2, 29.1, 62.4, 64.8, 128.4, 129.6, 132.9, 133.0, and 164.1.

Example 12 1,4-Butanediol mono(4-iodobenzoic acid)Ester (ABD-0069) (41B)

The title compound was prepared from 4-iodobenzoyl chloride and1,4-butanediol using Method 1 to give a clear oil (yield 90%). δ_(C)(CDCl₃, 62.9 MHz): 25.2, 29.2, 62.3, 65.1, 100.7, 129.8, 131.0, 137.7and 166.2.

Example 13 1,4-Butanediol mono(pentafluorobenzoic acid)Ester (ABD-0077)(4FB)

The title compound was prepared from pentafluorobenzoyl chloride and1,4-butanediol using Method 1 to give a clear oil. Column chromatographygave the title compound as the second fraction (yield 55%). δ_(C)(CDCl₃, 62.9 MHz): 25.0, 28.9, 62.2 and 66.7. δ_(H) (CDCl₃, 250 MHz):1.46 (1H, s, OH), 1.72 (2H, m), 1.86 (2H, m), 3.73 (2H, t, J 6.4) and4.42 (2H, t, J 6.4).

Example 14 1,4-Butanediol mono(2,3,6-trifluorobenzoic acid)Ester(ABD-0106) (2,3,6-FB)

The title compound was prepared from 2,3,6-trifluorobenzoic acid and1,4-butanediol using Method 2, to give a clear oil. Columnchromatography gave the title compound as the second fraction (yield60%). δ_(C) (CDCl₃, 62.9 MHz): 25.0, 29.0, 62.3, 66.2, 111.6 and 119.6.

Example 15 1,4-Butanediol mono(3,4-difluorobenzoic acid)Ester (ABD-0107)(3,4-FB)

The title compound was prepared from 3,4-difluorobenzoyl chloride and1,4-butanediol using Method 1, to give a clear oil. Columnchromatography gave the title compound as the second fraction (yield70%). δ_(C) (CDCl₃, 62.9 MHz): 25.2, 29.1, 62.3, 65.4, 117.4 (d, J17.6), 118.9 (d, J 18.6), 126.5 (d, J 3.9) and 127.3.

Example 16 1,4-Butanediol mono(2,3,4-trifluorobenzoic acid)Ester(ABD-0108) (2,3,4-FB)

The title compound was prepared from 2,3,4-trifluorobenzoic acid and1,4-butanediol using Method 2, to give a clear oil. Columnchromatography gave the title compound as the second fraction (yield85%). δ_(C) (CDCl₃, 62.9 MHz): 25.1, 29.1, 62.3, 65.7, 112.3 and 126.1.

Example 17 1,4-Butanediol mono(2,4,5-trifluorobenzoic acid)Ester(ABD-0109) (2,4,5-FB)

The title compound was prepared from 2,4,5-trifluorobenzoyl chloride and1,4-butanediol using Method 1, to give a white solid. Columnchromatography gave the title compound as the second fraction (yield65%). δ_(C) (CDCl₃, 62.9 MHz): 25.1, 29.0, 62.1, 65.7, 107.1 and 120.0.

Example 18 1,4-Butanediol mono(2,4-difluorobenzoic acid)Ester (ABD-0110)(2,4-FB)

The title compound was prepared from 2,4-difluorobenzoyl chloride and1,4-butanediol using Method 1, to give a clear oil. Columnchromatography gave the title compound as the second fraction (yield70%). δ_(C) (CDCl₃, 62.9 MHz): 25.1, 29.2, 62.3, 65.3, 105.2 (t, J26.4),111.6 (dd, J21.5, 2.9) and 133.9 (d, J 10.7).

Example 19 1,3-Propanediol mono[2-(4-isobutylphenyl)Propionic acid]ester(ABD-0037) (31)

The title compound was prepared from ibuprofen and 1,3-propanediol usingMethod 2 to give a clear oil (yield 70%). δ_(C) (CDCl₃, 62.9 MHz): 18.4,22.4, 30.2, 31.7, 45.0, 45.2, 59.1, 61.7, 127.1, 129.4, 137.7, 140.7 and175.3.

Example 20 1,4-Butanediol mono[2-(4-isobutylphenyl)Propionic acid]ester(ABD-0036) (41)

The title compound was prepared from ibuprofen and 1,4-butanediol usingMethod 2 to give a clear oil (yield 75%). δ_(C) (CDCl₃, 62.9 MHz): 18.4,22.4, 25.0, 29.0, 30.2, 45.0, 62.2, 64.5, 127.2, 129.3, 137.8, 140.6 and174.9.

Example 21 1,5-Pentanediol mono[2-(4-isobutylphenyl)Propionic acid]ester(ABD-0038) (51)

The title compound was prepared from ibuprofen and 1,5-pentanediol usingMethod 2 to give a clear oil (yield 70%). δ_(C) (CDCl₃, 62.9 MHz): 18.5,22.1, 22.4, 28.3, 30.2, 32.2, 45.0, 45.2, 62.7, 64.6, 127.2, 129.3,137.9, 140.5 and 174.9.

Example 22 1,6-Hexanediol mono[2-(4-isobutylphenyl)Propionic acid]ester(ABD-0039) (61)

The title compound was prepared from ibuprofen and 1,6-hexanediol usingMethod 2 to give a clear oil (yield 75%). δ_(C) (CDCl₃, 62.9 MHz): 18.5,22.4, 25.3, 25.6, 28.5, 30.2, 32.5, 45.0, 45.2, 62.7, 64.6, 127.2,129.3, 137.9, 140.5 and 174.9.

Example 23 1,4-Butanediol mono(4-benzylbenzoic acid)Ester (ABD-0034)(4PT)

The title compound was prepared from 4-benzylbenzoic acid and1,4-butanediol using Method 2, to give a clear oil (yield 45%). δ_(C)(CDCl₃, 62.9 MHz): 25.1, 29.1, 39.7, 62.4, 64.8, 126.0, 128.9, 129.1,131.7, 132.9, 130.1, 140.8, 143.4 and 172.8.

Example 24 1,4-Butanediol (biphenyl-2-carboxylic acid) ester (ABD-0059)(4BPX)

The title compound was prepared from biphenyl-2-carboxylic acid and1,4-butanediol using Method 2 to give a clear oil (yield 40%). δ_(C)(CDCl₃, 62.9 MHz): 24.6, 28.9, 62.3, 64.9, 127.3, 128.1, 128.5, 129.9,130.6, 130.7, 131.2, 131.9, 141.2, 141.7, 142.3, 143.2 and 169.2.

Example 25 1,3-Propanediol mono(biphenyl-4-carboxylic acid)Ester(ABD-0057) (3BP)

The title compound was prepared from biphenyl-4-carbonyl chloride and1,3-propanediol using Method 1 to give a white solid (yield 80%). δ_(C)(CDCl₃, 62.9 MHz): 14.2, 30.9, 32.0, 127.1, 127.3, 128.2, 129.3, 130.1,140.0, 145.8 and 166.9.

Example 26 1,4-Butanediol mono(biphenyl-4-carboxylic acid)Ester(ABD-0056) (4BP)

The title compound was prepared from biphenyl-4-carbonyl chloride and1,4-butanediol using Method 1 to give a white solid (yield 85%). δ_(C)(CDCl₃, 62.9 MHz): 25.3, 29.3, 62.4, 64.8, 127.1, 127.3, 128.2, 129.0,129.1, 130.1, 130.7, 140.0, 145.7 and 166.6. δ_(H) (CDCl₃, 250 MHz):1.74-1.77 (2H, m), 1.86-1.89 (2H, m), 2.63 (1H, s, OH), 3.74 (2H, t,J6.3), 4.38 (2H, t, J6.3), 7.45 (3H, m), 7.63 (4H, m) and 8.10 (2H, d, J8.5). m/z (Found M, 270. C₁₇H₁₈O₃ requires 270).

Example 27 1,5-Pentanediol mono(biphenyl-4-carboxylic acid)Ester(ABD-0055) (5BP)

The title compound was prepared from biphenyl-4-carbonyl chloride and1,5-pentanediol using Method 1 to give a white solid (yield 80%). δ_(C)(CDCl₃, 62.9 MHz) 22.4, 28.6, 32.4, 62.7, 65.0, 127.1, 127.3, 128.2,129.0, 129.1, 130.1, 140.0, 145.6 and 166.7; m/z (Found M, 284.C₁₈H₂₀O₃requires 284).

Example 28 1,6-Hexanediol mono(biphenyl-4-carboxylic acid)Ester(ABD-0054) (6BP)

The title compound was prepared from biphenyl-4-carbonyl chloride and1,6-hexanediol using Method 1 to give a white solid (yield 85%). δ_(C)(CDCl₃, 62.9 MHz) 25.5, 25.9, 28.8, 32.7, 62.9, 65.0, 127.1, 127.3,128.2, 129.0, 129.2, 130.1, 140.1, 145.7 and 166.7. m/z (Found M, 298.C₁₉H₂₂O₃ requires 298).

Example 29 2,2,3,3-Tetrafluoro-butane-1,4-diolmono(biphenyl-4-carboxylic acid)Ester (ABD-0095) (BP-4F)

The title compound was prepared from biphenyl-4-carbonyl chloride and2,2,3,3-tetrafluorobutane-1,4-diol using Method 1 to give a clear oil(yield 65%). Purification by column chromatography (ethyl acetate:lightpetroleum, 1:1) gave the title compound as the second fraction. δ_(C)(CDCl₃, 62.9 MHz): 60.4 (t, J 26.4), 127.3, 127.3, 128.4, 129.0, 130.6,139.8, 146.5 and 165.4.

Example 30 1,4-Butanediol mono(4′-methyl-biphenyl-4-carboxylicacid)Ester (ABD-0070) (Me4BP)

The title compound was prepared from 4′-methyl-biphenyl-4-carboxylicacid (prepared from 4-bromotoluene using Method 6) and 1,4-butanediolusing Method 2 to give a white solid (yield 20%). δ_(C) (CDCl₃, 62.9MHz): 21.2, 25.3, 29.3, 62.4, 64.8, 126.8, 127.1, 128.8, 129.7, 130.1,137.1, 138.2, 145.6 and 166.7.

Example 31 1,4-Butanediol mono(4′-hydroxy-biphenyl-4-carboxylicacid)Ester (ABD-0072) (HO4BP)

The title compound was prepared from 4′-hydroxy-biphenyl-4-carboxylicacid (which was first protected as the4′-tertiarybutylchlorodiphenylsilyl ether) and 1,4-butanediol usingMethod 2 (the silyl protecting group was removed during the reaction) togive the title product as a white powder (yield 15%). δ_(C) (CDCl₃, 62.9MHz): 25.1, 29.0, 60.3, 64.6, 115.9, 126.0, 127.6, 128.2, 129.5, 144.7,158.0 and 165.7.

The protected acid was prepared as follows:4′-Hydroxy-biphenyl-4-carboxylic acid (4.2 g, 20 mmol) was dissolved inpyridine (50 ml). Tertiarybutylchlorodiphenylsilane (TBDPSi—Cl) (11 g,40 mmol) was added dropwise, followed by a catalytic amount of4-dimethylaminopyridine (0.1 g). The solution was left to stirovernight, then poured into water (200 ml) and extracted with methylenechloride (100 ml). The organic phase was washed with 2 M HCl (100 ml)which caused the precipitation of an acid. The acid was collected byfiltration, dissolved in diethyl ether and washed with water. Themethylene chloride layer was washed with water, the two organic phasescombined and dried over Na₂SO₄. Evaporation and purification by columnchromatography (light petroleum:ethyl acetate, 2:1) gave the silylatedproduct as a white solid.

Example 32 1,4-Butanediol mono(3′,4′-dimethyl-biphenyl-4-carboxylicacid)Ester (ABD-0089) (Xy4BP)

The title compound was prepared from3′,4′-dimethyl-biphenyl-4-carboxylic acid (prepared from4-bromo-o-xylene using Method 6) and 1,4-butanediol using Method 2 togive a clear oil (yield 15%). δ_(C) (CDCl₃, 62.9 MHz): 19.5, 20.0, 25.3,29.2, 62.3, 64:8; 124.6, 126.8, 128.5, 128.6, 130.0, 130.2, 136.8,137.2, 137.5, 145.8 and 166.8.

Example 33 1,4-Butanediol mono(4′-ethyl-biphenyl-4-carboxylic acid)Ester(ABD-0094) (Et4BP)

The title compound was prepared from 4′-ethyl-biphenyl-4-carboxylic acid(prepared from 4-bromo-ethylbenzene using Method 6) and 1,4-butanediolusing Method 2 to give a white powder (yield 25%). δ_(C) (CDCl₃, 62.9MHz): 15.6, 25.3, 28.6, 29.3, 62.4, 64.8, 126.9, 127.2, 128.5, 128.8,130.1, 137.3, 144.5, 145.6 and 166.7. δ_(H) (CDCl₃, 250 MHz): 1.27 (3H,t, J 7.6), 1.76 (2H, m), 1.86 (2H, m), 2.43 (1H, br s), 2.70 (2H, q, J7.6), 3.74 (2H, t, J 6.4), 4.38 (2H, t, J 6.4), 7.29 (2H, d, J 8.2),7.54 (2H, d, J 8.2), 7.64 (2H, d, J 8.2), 8.08 (2H, d, J 8.2).

Example 34 1,4-Butanediol mono(4′-methoxy-biphenyl-4-carboxylicacid)Ester (ABD-0097) (4-OMeBP)

The title compound was prepared from 4′-methoxy-biphenyl-4-carboxylicacid (prepared from 4-methoxybiphenyl using Methods 4 and 5) and1,4-butanediol using Method 2. Purification by column chromatography(ethyl acetate:light petroleum 1:1) gave the title compound as a whitepowder (yield 45%). δ_(C) (CDCl₃, 62.9 MHz): 25.3, 29.3, 55.4, 62.3,64.8, 114.4, 126.5, 128.4, 130.1, 132.4, 145.2, 159.8 and 166.7. δ_(H)(CDCl₃, 250 MHz): 1.60 (1H, s, OH), 1.76 (2H, m), 1.85 (2H, m), 3.73(2H, t, J 6.4), 3.85 (3H, s, OMe), 4.37 (2H, t, J 6.4), 6.98 (2H, d, J8.8), 7.56 (2H, d, J 8.8), 7.61 (2H, d, J 8.2) and 8.06 (2H, d, J 8.5).

Example 35 1,4-Butanediol mono(2′-nitro-biphenyl-4-carboxylic acid)Ester(ABD-0098) (2-NO₂BP)

The title compound was prepared from 2′-nitro-biphenyl-4-carboxylic acid(prepared from 2-nitro-biphenyl using Methods 4 and 5) and1,4-butanediol using Method 2. Purification by column chromatography(ethyl acetate:light petroleum, 1:1) gave the title compound as a paleyellow oil (yield 15%). δ_(C) (CDCl₃, 62.9 MHz): 25.3, 29.2, 62.4, 65.0,124.4, 128.0, 128.9, 129.9, 130.1, 131.8, 132.7, 135.6, 142.2, 149.0 and166.2. δ_(H) (CDCl₃, 250 MHz): 1.62 (1H, s, OH), 1.76 (2H, m), 1.85 (2H,m), 3.73 (2H, t, J 6.4), 4.38 (2H, t, J 6.4), 7.37 (2H, d, J 8.5), 7.41(1H, dd, J 7.9, 1.5), 7.52 (1H, J 7.9, 1.5), 7.64 (1H, td, J7.6, 1.2),7.91 (1H, d, 8.2) and 8.10 (2H, d, J 8.5).

Example 36 1,4-Butanediol mono(2′-fluoro-biphenyl-4-carboxylicacid)Ester (ABD-0099) (2-FBP)

The title compound was prepared from 2′-fluoro-biphenyl-4-carboxylicacid (prepared from 2-fluorobiphenyl using Methods 4 and 5) and1,4-butanediol using Method 2. Purification by column chromatography(ethyl acetate:light petroleum, 1:1) gave the product as a clear oil(yield 25%). δ_(C) (CDCl₃, 62.9 MHz): 25.3, 29.2, 62.4, 64.9, 116.3 (d,J23.4), 124.5, (d, J2.9), 128.0 (d, J 13.7), 129.0 (d, J 2.0), 129.4,129.7, 129.8 (d, J 13.7), 130.6 (d, J 2.0), 140.4, 159.6 (d, J 249.0)and 166.6. δ_(H) (CDCl₃): 1.67 (1H, s, OH), 1.76 (2H, m), 1.88 (2H, m),3.73 (2H, t, J 6.4), 4.38 (2H, t, J 6.4), 7.16 (1H, m), 7.24 (1H, dd J8.8, 1.2), 7.33 (1H, m), 7.44 (1H, dt, J 7.6, 1.8) 7.61 (2H, dd, J 8.5,1.8) and 8.10 (2H, d, J 8.5).

Example 37 1,4-Butandiol mono(4′-fluoro-biphenyl-4-carboxylic acid)Ester(ABD-0100) (4-FBP)

The title compound was prepared from 4′-fluoro-biphenyl-4-carboxylicacid (prepared from 4-fluorobiphenyl using Methods 4 and 5) and1,4-butanediol using Method 2. Purification by column chromatography(ethyl acetate:light petroleum, 1:1) gave the title compound as a whitesolid (yield 50%). δ_(C) (CDCl₃, 62.9 MHz): 25.3, 29.2, 62.4, 64.9,115.9 (d, J22.5), 126.9, 128.9 (d, J8.8), 129.1, 130.2, 136.1 (d, J2.9),144.6, 163.0 (d, J 248.0) and 166.6. δ_(H) (CDCl₃, 250 MHz): 1.58 (1H,s, OH), 1.74 (2H, m), 1.86 (2H, m), 3.74 (2H, t, J 6.4), 4.38 (2H, t, J6.4), 7.14 (2H, t, J 8.8), 7.56 (2H, d, J 8.8), 7.59 (2H, dd, J 8.2) and8.08 (2H, d, J 8.5).

Example 38 1,4-Butanediol mono(4′-bromo-biphenyl-4-carboxylic acid)Ester(ABD-0102) (4-BrBP)

The title compound was prepared from 4′-bromo-biphenyl-4-carboxylic acid(prepared from 4-bromobiphenyl using Methods 4 and 5) and 1,4-butanediolusing Method 2. Purification by column chromatography (ethylacetate:light petroleum, 1:1) gave the title compound as a white solid(yield 30%). δ_(C) (DMSO, 62.9 MHz): 25.3, 29.3, 62.4, 64.9, 122.6,126.9, 128.9, 129.4, 130.2, 132.1, 138.0, 144.4 and 166.4.

Example 39 1,4-Butandiol mono(triphenylacetic acid)Ester (ABD-0028) (4T)

The title compound was prepared from triphenylacetyl chloride (preparedby from triphenylacetic acid using Method 8) and 1,4-butanediol usingMethod 1, to give the title compound as a white powder (yield 45%).δ_(C) (CDCl₃, 62.9 MHz): 24.8, 29.0, 62.2, 65.5, 67.6, 126.9, 127.8,130.3, 143.0 and 173.7.

Example 40 1,5-Pentandiol mono(triphenylacetic acid)Ester (ABD-0030)(5T)

The title compound was prepared from triphenylacetyl chloride (preparedby from triphenylacetic acid using Method 8) and 1,5-pentanediol usingMethod 1, to give the title compound as a white powder (yield 35%).δ_(C) (CDCl₃, 62.9 MHz): 22.1, 28.1, 32.1, 62.6, 65.6, 67.6, 126.9,127.7, 130.3, 143.0 and 173.7.

Example 41 1,6-Hexandiol mono(triphenylacetic acid)Ester (ABD-0031) (6T)

The title compound was prepared from triphenylacetyl chloride (preparedby from triphenylacetic acid using Method 8) and 1,6-Hexanediol usingMethod 1, to give the title compound as a white powder (yield 35%).δ_(C) (CDCl₃, 62.9 MHz): 25.2, 25.6, 28.3, 32.5, 62.8, 65.6, 67.6,126.9, 127.7, 130.3, 143.0 and 173.7.

Example 42 1,3-Propanediol mono(biphenyl-4-yl-acetic acid)Ester(ABD-0041) (3BPA)

The title compound was prepared from biphenyl-4-yl-acetic acid and1,3-propanediol using Method 2 to give a white solid (yield 75%). δ_(C)(CDCl₃, 62.9 MHz): 31.7, 41.1, 59.1, 62.0, 127.1, 127.4, 128.8, 129.7,133.0, 140.2, 140.8 and 172.1.

Example 43 1,4-Butanediol mono(biphenyl-4-yl-acetic acid)Ester(ABD-0042) (4BPA)

The title compound was prepared from biphenyl-4-yl-acetic acid and1,4-butanediol using Method 2 to give a white solid (yield 80%). δ_(C)(CDCl₃, 62.9 MHz): 25.1, 29.1, 41.1, 62.3, 64.8, 127.1, 127.4, 128.8,129.7, 133.1, 140.1, 140.8 and 171.8. m/z (Found M, 284.C₁₈H₂₀O₃requires 284).

Example 44 1,5-Pentanediol mono(biphenyl-4-yl-acetic acid)Ester(ABD-0043) (5BPA)

The title compound was prepared from biphenyl-4-yl-acetic acid and1,5-pentanediol using Method 2 to give a white solid (yield 75%). δ_(C)(CDCl₃, 62.9 MHz): 22.2, 28.4, 32.3, 41.1, 62.6, 65.0, 127.1, 127.3,128.8, 129.7, 133.2, 140.1, 140.8 and 171.8.

Example 45 1,6-Hexanediol mono(biphenyl-4-yl-acetic acid)Ester(ABD-0044) (6BPA)

The title compound was prepared from biphenyl-4-yl-acetic acid and1,6-hexanediol using Method 2 to give a white solid (yield 70%). δ_(C)(CDCl₃, 62.9 MHz): 25.4, 25.7, 28.6, 32.6, 41.1, 62.8, 65.0, 127.1,127.3, 128.8, 129.7, 133.2, 140.0, 140.8 and 171.8.

Example 46 1,4-Butanediol mono(naphth-1-yl-acetic acid)Ester (ABD-0032)(4N)

The title compound was prepared from naphth-1-yl acetic acid and1,4-butanediol using Method 2, to give a clear oil (yield 70%). δ_(C)(CDCl₃): 25.0, 29.0, 39.3, 62.1, 64.8, 123.8, 125.6, 125.8, 126.4,128.1, 128.1, 128.9, 130.7, 132.1, 133.8 and 171.8.

Example 47 1,4-Butanediol mono(homoveratric acid)Ester (ABD-0033) (4H)

The title compound was prepared from homoveratric acid and1,4-butanediol using Method 2 to give a clear oil (yield 90%). δ_(C)(CDCl₃, 62.9 MHz): 25.1, 29.0, 41.0, 55.9, 62.2, 64.7, 111.2, 112.4,121.4, 126.5, 148.1, 148.9 and 172.0.

Example 48 Butyl [2-(4-isobutylphenyl)Propionic acid] ester (ABD-0035)(Bul)

The title compound was prepared from ibuprofen and butanol using Method2 to give a clear oil (yield 85%). δ_(C) (CDCl₃, 62.9 MHz): 13.7, 18.5,19.0, 22.4, 30.2, 30.6, 45.1, 45.2, 64.6, 127.2, 129.3, 137.9, 140.5 and174.9.

Example 49 Butyl (biphenyl-4-yl-acetic acid) ester (ABD-0040) (BuBPA)

The title compound was prepared from biphenylacetic acid and butanolusing Method 2 to give a clear oil (yield 70%). δ_(C) (CDCl₃, 62.9 MHz):13.8, 19.1, 30.7, 41.1, 64.9, 127.1, 127.3, 128.8, 129.7, 133.3, 140.0,140.9 and 171.7.

Example 50 Butyl (biphenyl-4-carboxylic acid) ester (ABD-0053) (BUBP)

The title compound was prepared from biphenyl-4-carbonyl chloride andbutanol using Method 1 to give a white solid (yield 85%). δ_(C) (CDCl₃,62.9 MHz) 13.8, 19.3, 30.9, 64.9, 127.1, 127.3, 128.1, 129.0, 130.1,140.1, 145.6 and 166.6; m/z (Found M, 254. C₁₇H₁₈O₂ requires 254).

Example 51 Pentyl (biphenyl-4-carboxylic acid) ester (ABD-0090) (PBP)

The title compound was prepared from biphenyl-4-carbonyl chloride andpentanol using Method 1 to give a clear oil (yield 85%). δ_(C) (CDCl₃,62.9 MHz): 14.1, 22.4, 28.3, 28.5, 65.2, 127.1, 127.3, 128.1, 128.9,129.3, 130.1, 140.1, 145.6 and 166.6.

Example 52 4-Methoxybutyl (biphenyl-4-carboxylic acid) ester (ABD-0050)(4BP-OMe)

ABD-0056 (7 mmol) was dissolved in acetone (20 ml) containing powderedNaOH (1.5 g). Dimethyl sulphate (1.5 g, 12 mmol) was added dropwise andthe mixture stirred overnight to give a slurry. The slurry was pouredinto water (200 ml) and washed with methylene chloride (100 ml). Theorganic phase was washed with water until it became clear. The organicphase was dried over Na₂SO₄, the solvent evaporated and the titlecompound obtained by column chromatography (light petroleum:ethylacetate, 1:1, followed by a repeat using light petroleum:ethyl acetate,5:1) as a yellow oil (yield 50%). δ_(C) (CDCl₃, 62.9 MHz): 25.6, 26.3,58.7, 64.8, 72.2, 127.1, 127.3, 128.2, 129.0, 129.2, 130.1, 140.0, 145.6and 166.6.

Example 53 4-Bromobutyl (biphenyl-4-carboxylic acid) ester (ABD-0086)(4BP-Br)

The title compound was prepared from biphenyl-4-carbonyl chloride and4-bromobutanol (prepared using Method 9) using Method 1, to give thetitle compound as a pale brown oil (yield 20%). δ_(C) (CDCl₃, 62.9 MHz):26.2, 29.3, 44.6, 64.2, 127.1, 127.3, 128.2, 129.0, 130.1, 140.0, 145.7and 166.5.

Example 54 4-Nitrooxybutyl (biphenyl-4-carboxylic acid) ester (ABD-0087)(4BP-NO₂)

The title compound was prepared from ABD-0086 (5 mmol) by stirring withAgNO₃ (25 mmol) in acetonitrile (50 ml) for 24 hours. The mixture wasfiltered, the filtrate evaporated and purified by column chromatography(light petroleum:ethyl acetate, 4:1) to give the title compound as paleyellow oil (yield 55%). δ_(C) (CDCl₃, 62.9 MHz): 23.8, 25.2, 64.0, 72.7,127.1, 127.3, 128.2, 128.8, 129.0, 130.1, 139.9, 145.8 and 166.4.

Example 55 4-Nitrooxybutyl (2,2′,4′-trinitro-biphenyl-4-carboxylic acid)ester (ABD-0088) (4×NO₂-BP)

ABD-0056 (10 mmol) was stirred in nitric acid (50 ml) and thetemperature slowly increased to 80° C. for 5 hours. The mixture waspoured into water (250 ml) and extracted with methylene chloride (100ml). The organic phase was washed with water, saturated NaHCO₃ andwater, dried with Na₂SO₄ and evaporated. Column chromatography (ethylacetate:light petroleum, 1:1) gave the title compound as a thick yellowoil (yield 20%). δ_(C) (CDCl₃, 62.9 MHz): 23.7, 25.1, 65.3, 72.4, 120.6,126.2, 127.9, 130.7, 131.9, 132.4, 134.5, 136.5, 139.7, 146.7, 147.0,148.0 and 163.7. δ_(H) (CDCl₃, 250 MHz): 1.95 (4H, m), 4.47 (2H, m),4.54 (2H, m), 7.40 (1H, d, J7.9), 7.54 (1H, d, J8.5), 8.38 (1H, dd,J7.9, 1.8), 8.55 (1H, dd, J8.5, 2.4), 8.90 (1H, d, J 1.5) and 9.10 (1H,d, J2.4).

Biological Studies

Initial screening of candidate compounds was performed using viabilityassays, on cultures of the macrophage cell line J774, which have beenused before as a model system for osteoclast survival (see, e.g.,Luckman et al., 1998). The assay is based on the survival of the J774macrophage cell line; macrophages are closely related to osteoclasts,and contain similar high levels of esterase activity.

MTT Macrophage J774 Viability Assay

J774 cells were plated at 10⁴ cells per well in 150 μL αMEM (α ModifiedEagle Medium) in 96-well plates and grown overnight. The next day,compounds were added to the cultures, and culture was continued foranother 72 hours. At the end of the culture period cell survival wasdetermined using the tetrazolium dye-based MTT assay as previouslydescribed (see, e.g., MacPherson et al., 1999).

MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) hasan orange colour and is soluble in the medium used for cell culture. Themitochondrial enzyme succinate dehydrogenase acts upon MTT in livingcells to produce the insoluble purple coloured formazan. The amount offormazan produced, as measured by UV/visible spectroscopy, isproportional to the number of viable cells.

Briefly, MTT (5 mg/ml MTT in αMEM) was added to each well (1:10 v/v, 15μL) and the cells incubated for 4 hours. The medium was carefullyremoved using a needle without dislodging the crystal layer. 100 μLacidified isopropanol (4 M HCl 1:100 v/v in isopropanol) was added toeach well and the purple crystals allowed to dissolve. The absorbancewas measured in a plate reader at 540 nm, with 690 nm as reference. Thecontrols were a deep purple colour, indicating a high number of livecells. The results for each test compound were expressed as a % of theaverage control value.

Addition of Compounds. All of the compounds studied were made up as 100mM solutions in DMSO. These stock solutions were then diluted 100× inculture medium. From these 1 mM solutions, convenient quantities (3-15μL) were added directly to the wells so as to give the desired finalcompound concentration.

Alamar Blue Macrophage J774 Viability Assay

J774 cells were plated at 10⁴ cells per well in 150 μL αMEM (α ModifiedEagle Medium) in 96-well plates and grown overnight. The next day,compounds were added to the cultures, and culture was continued foranother 72 hours. At the end of the culture period cell survival wasdetermined using an Alamar Blue assay as previously described (see,e.g., Nociari et al., 1998).

Alamar Blue is an oxidation-reduction sensitive indicator. The dyeitself is in the oxidised state, which is blue and non-fluorescent. Thedye can accept electrons from reducing species, such as NADPH and FADH,to form a reduced dye species, which is red and fluorescent. Thus thetransformation from oxidised form to reduced form can be measured byfluorimetric or colourimetric means. For fluorescence measurements,530-560 nm excitation and 590 nm emission wavelengths are typicallyused. For colourimetric measurements, absorbance is measured at 570 nm(reduced form) and 600 nm (oxidised form) and a simple calculationperformed to determine the relative quantities of the two species.

A high ratio of the reducing species, NADPH and FADH, to thecorresponding oxidised species, NADP and FAD, is an indicator that cellsare proliferating and viable. A low ratio indicates cells that arequiescent or non-viable.

Briefly, Alamar Blue (Biosource International) was added undiluted tothe each well (1:10 v/v, 15 μL). The plate was incubated at 37° C. for3-4 hours and the fluorescence was measured at 570 nm, with a 25 nmbandwidth. A high reading indicated cells with normal viability, and alow reading indicates cells that have been damaged and are no longerproliferating normally. The controls gave a high fluorescence reading,indicating a high number of live, healthy cells. A potent test compoundgave a low fluorescence reading. The average results for each testcompound (n=5) were expressed as a % of the average control value.

Addition of Compounds. All of the compounds studied were made up as 100mM solutions in DMSO. These stock solutions were then diluted 100 or1000× in culture medium (αMEM). From these 1 mM or 100 μM solutions,convenient quantities (3-15 μL) were added directly to the wells so asto give the desired final compound concentration.

This assay offers numerous advantages over other assays, including MTTassays: it permits a higher throughput; it is more sensitive; it isnon-damaging to the cells; it is faster; it generally gives an identicalresult to MTT assay. A comparison is shown in FIG. 1, which is a graphof the macrophage J774 viability, as measured by the MTT and Alamar Bluemacrophage J774 viability assays, expressed as % of control, after 72hours exposure to ABD-0028 and ABD-0042, as a function of concentrationof compound. Only one compound (ABD-0056) showed significant differencebetween the MTT and Alamar Blue assays.

Additional Studies

Some compounds were further evaluated in two model systems of trueosteoclasts: (a) the murine co-culture system, and (b) the rabbitosteoclast culture system.

Murine Co-Culture System

The first model system, the murine co-culture system, studies theformation of osteoclasts_from precursors present in the bone marrow. Thenumber of osteoclasts and the amount of dentine resorption was measured.

Osteoclast formation and activity was studied using an adaptation (see,e.g., van't Hof & Ralston, 1997) of the osteoblast-bone marrowco-culture assay originally described by Takahashi et al., 1988.

Co-Culture Methods. Co-culture (see, e.g., Van't Hof et al., 1997) is amethod to study the formation of osteoclasts from their precursors. Inthis assay, osteoblasts were obtained from the calvaria of 2-3 day oldneonatal mice. These were plated on dentine, stimulated with1,25-dihydroxy vitamin D₃ to stimulate RANKL and M-CSF expression. Earlyosteoclast precursors were present in the bone marrow of adult mice. Thebone marrow suspension was purified to remove the red blood cells andthe remainder cultured on top of the osteoblast layer. The stimulatoryfactors then allowed the osteoclast precursors to differentiate intomature osteoclasts. At the end of the culture osteoclasts wereidentified by TRAcP staining and the resorption activity was measured inthe same manner as for rabbit osteoclasts.

Although it is possible to generate osteoclasts from bone marrow cellsalone by treating the cultures with RANKL and M-CSF, the co-culturesystem is still regarded as one of the most reliable and reproducibleavailable. It is useful for studying the effects of drugs on bothosteoclast progenitors and mature osteoclasts.

Preparation of Dentine. The dentine was elephant ivory, preferred tobone because of its uniform surface, which facilitates easyvisualisation of resorption pits. It was cut into slices ofapproximately 200 μm thickness using a Buehler Isomet low speed saw witha diamond wafering blade (series 15 HC). These slices were polished byhand, to a high degree, until one side was shiny. Out of these slices,discs were punched that fit the wells of a 96 well plate, using a paperpuncher. Excess residues from the polish were removed by sonication. Thediscs were then stored in 70% ethanol until required. These discs werethen dried and placed shiny side up in the wells of a 96 well plate.Cells were seeded onto the dentine. Following completion of the culture,these dentine slices were carefully removed from the plate and studiedunder the microscope.

Osteoblast Isolation. Briefly, osteoblasts were isolated from thecalvarial bones of 2-day-old mice by sequential collagenase digestion(type I collagenase, Sigma) and cultured in αMEM supplemented with 10%FCS (foetal calf serum) and penicillin and streptomycin at 37° C. in 5%CO₂.

More specifically, osteoblasts were obtained from a collagenasedigestion of the calvaria (skull bones) of 2-3 day old neo-natal MF1mice. At this stage in their development these are soft and easilyremoved. The calvaria from 5-6 mice were carefully dissected and washedin HBSS (Hank's balanced saline solution). The calvaria were placed in a15 ml tube and shaken at 37° C. in 4 ml collagenase (10 mg/ml) for 10minutes. This removes the excess unwanted tissue. The liquid wasdisposed of and a further 4 ml collagenase (10 mg/ml) added to the tube.The calvaria were then digested for a further 30 minutes. After this thesupernatant (F1) was removed and retained. The calvaria were washed witha 2×4 ml PBS and this was added to F1. 4 ml EDTA (ethylene diaminetetraacetic acid) (4 mM in PBS) was then added to complex the calciumand allow further extraction of osteoblasts. This was shaken for 10minutes at 37° C. The supernatant was removed and retained (F2). Thecalvaria were again washed with 2×4 ml HBSS and this was added to F2.The final 4 ml of collagenase (10 mg/ml) was added to the tube and thiswas again shaken at 37° C. for 30 minutes. Whilst this was being done,F1 and F2 were spun down at 300 g for 3 minutes, brake 3. The pelletswere re-suspended in 1 ml medium (αMEM supplemented with 10% FCS (foetalcalf serum) and penicillin and streptomycin), combined and added to 10ml medium in two 75 cm² flasks. The liquid from the final collagenasedigestion was collected (F3), the calvaria washed and the combinedliquid extracts spun down in the centrifuge. The pellet was re-suspendedin 1 ml medium and added in equal proportions to the flasks containingF1 and F2. The flasks were left for 4-6 hours at 37° C. and then themedium was changed to remove any non-adherent cells. These flasks may beleft for up to 4 days at 37° C., 5% CO₂.

Osteoblast Plating. The medium was removed from the flasks and the cellswashed with PBS. 2 ml Trypsin was added to the cells and these wereincubated at 37° C. for 2 minutes. The flasks usually required gentleagitation to fully loosen the cells. 4 ml medium supplemented with 10%FCS was added to stop the enzymatic action. The cells were removed andthe flask washed out with medium. The cell suspension was spun down at300 g for 3 minutes, the medium removed and the pellet re-suspended in 1ml medium. The cells were counted and then seeded in a 96 well platedcontaining dentine slices, at 8×10³ cells per well in 100 μl mediumcontaining 1000× dilution of stock 1,25-dihydroxyvitamin D₃ (final conc.10 nM/well) to stimulate the expression of RANKL and cultured overnight.

Isolation of Bone Marrow Cells. Briefly, bone marrow cell populationscontaining osteoclast precursors were isolated from the long bones of3-5 month old mice and erythrocytes were removed by Ficoll Hypaquedensity gradient centrifugation. The resulting bone marrow cells werewashed with PBS (phosphate buffered saline) and re-suspended in culturemedium.

More specifically, the femurs and tibia were dissected from 2-3 adultMF1 mice 3-6 months old) and the surrounding tissue was removed. Thebones were trimmed to allow access to the bone marrow. The marrow wasflushed out using a 25G needle and HBSS+10% FCS. A single cellsuspension is obtained by repeatedly squeezing the cell suspensionthrough needles of decreasing size (start with 19G, end with 25G). 5 mlFicoll was added to a 15 ml tube and the cell suspension carefullyplaced on top of this with the minimum amount of mixing between thelayers. The density centrifugation was performed at 600 g, 25 min, brakeoff. This was sufficient to allow the red blood cells to congregate atthe bottom of the tube, fats to remain at the top of the liquid and thedesired bone marrow cells to collect at the interface. The cloudy layerfrom the interface was collected with a pipette, placed in a fresh 15 mltube and made up to 12 ml with HBSS. The cell suspension was spun downat 300 g for 3 minutes. The pellet was collected and re-suspended in 1ml medium. The bone marrow cells were counted and then added to the 96well plate containing the osteoblasts at 2×10⁵ cells/well in 50 μLmedium.

Osteoblast Precursor Studies. To investigate the effects of a drug onosteoclast precursors the timetable was as follows:

-   Day 0—Plate osteoblasts.-   Day 1—Plate bone marrow cells-   Day 2—Add test compound.-   Day 4—100% medium refresh+1,25-dihydroxyvitamin D₃(final conc. 10    nm/well)-   Day 6—Add IL1 (10 u/ml) and 1,25-dihydroxyvitamin D₃ (final conc. 10    nm/well)-   Day 10—Fix cells.

Mature Osteoclast Studies. To investigate the effects of a drug onmature osteoclasts the timetable was as follows:

-   Day 0—Plate osteoblasts.-   Day 1—Plate bone marrow cells.-   Day 6—50% medium refresh+10 nM IL1 and 1,25-dihydroxyvitamin D₃.-   Day 7—Add drugs and remove and fix day 7 control slices.-   Day 10—Fix cells.

At the conclusion of a study, the cells were fixed in 4% formaldehydefor 10 minutes and washed in PBS. Fixed cells were either stained andkept in 70% ethanol or refrigerated in water or PBS. The 50% mediumrefresh involved the addition of 150 μL fresh medium containing a 500×dilution of 1,25-dihydroxyvitamin D₃ and a 250× dilution of IL1(interleukin 1). This was left for 15 minutes and then 150 μL mediumcarefully removed. The medium refresh must be done very carefully,because the confluent layer of osteoblasts can be quite easilydisturbed, and detached. This would result in a total absence ofosteoclasts. Usually the first osteoclasts and resorption pits appearedon day 6. Reasonable numbers of osteoclasts were present between day7-10.

At the end of the culture, the osteoclasts were identified by stainingfor tartrate-resistant acid phosphatase (TRAcP) staining and resorptionpit area was quantified by reflected light microscopy as describedpreviously (see, e.g., van't Hof & Ralston, 1997).

TRAcP Staining. Osteoclasts express very high levels of the enzymetartrate resistant acid phosphatase (TRAcP) and can therefore be easilyvisualised by staining for this enzyme, for example, by the followingmethod. Two staining solutions, (1) and (2), were made up freshly asfollows:

-   Solution 1. 300 μL Naphthol-AS-BI-phosphate stock.    -   1.5 ml Veronal buffer.    -   1.8 ml Acetate buffer.    -   1.8 ml Acetate buffer with 100 mM tartrate.-   Solution 2. 240 μL Pararosaniline.    -   240 μL NaNO₂ (4% stock solution).

Naphthol-AS-BI-phosphate stock: 10 mg/ml Naphthol-AS-BI-phosphate indimethylformamide.

Veronal buffer: 1.17 g anhydrous Sodium Acetate; 2.94 g Veronal (sodiumbarbiturate); dissolved in 100 ml distilled water.

Acetate buffer 0.1 M, pH 5.2: solution (a): 0.82 g Sodium Acetateanhydrous dissolved in 100 ml distilled water; solution (b): 0.6 mlAcetic acid glacial made up to 100 ml with distilled water; pH ofsolution (a) adjusted to pH 5.2 with solution (b).

Pararosaniline: 1 g Pararosaniline in 20 ml distilled water. 5 mlconcentrated hydrochloric acid was added, the solution was heatedcarefully in a water bath while stirring. The solution was allowed tocool and then filtered.

Solutions (1) and (2) were mixed and filtered to give the stainingsolution. The PBS from the wells was removed and at least 50 μL ofstaining solution added.

The cells were incubated at 37° C. for about 45 min, or until thedentine slices appeared sufficiently red. To determine what passes assufficient it was necessary to remove the dentine slice and check undera light microscope that the osteoclasts were suitably stained. Thestaining solution was then removed and replaced with 70% ethanol. Thedentine-slices-were stored in a refrigerator.

Osteoclast Counting. This was done using a light microscope to determinethe number of TRAcP positive multinucleated cells on each dentine slice.The slices were carefully removed from the 96-well plate, avoidingdisturbance of the cell layer, and placed on a glass slide. A few dropsof 70% ethanol were put on each slice followed by a glass coverslip.Working across the dentine the number of multinucleated, red-stainedcells were counted. There were usually a large number of small redmononucleated cells. These were osteoclast precursors and these were notcounted. The numbers of osteoclasts on the control slices can range from300 up to 1000. For each compound or concentration studied, the averageof the values for the 5 slices was taken and expressed as a % of theaverage value for the controls. Any obvious outlying values wereignored. The most common reason for this was when there were no cells ofany kind, usually indicating that the osteoblast layer has detachedduring handling.

Quantification of Resorption Area. After the osteoclasts were stainedand counted it was necessary for the dentine slice to be thoroughlycleaned. The slices were rubbed on a suitable surface, a piece of blueroll proved ideal for this purpose. In order to clean the slicesproperly it may be necessary to wash them in dilute HClO for a fewseconds to loosen the cell debris. The resorption pits can be visualisedeither by staining with dyes such as Toluidine blue or Coomassie blue,by scanning electron microscopy or by reflected light microscopy. Here,reflected light microscopy was used, because it is easy to perform, theslices needed only thorough cleaning and no staining, and the imageobtained could be fairly easily quantified using image analysis. Becausethe slices need to be completely flat for the reflected lightmicroscopy, they were glued glass slides under pressure of a 0.5 kgmetal weight. These may then be easily stored. A Zeiss reflected lightmicroscope was used, fitted with a 2.5× lens, wide field c-mountadapter, and Diagnostics Instruments Insight BNV large chip digitalcamera. This set-up allowed the capture of an entire bone slice in oneimage at sufficient resolution to identify and measure the resorptionpits. The image analysis software package was developed using theAphelion ActiveX image analysis toolkit from ADCIS (ADCIS SA,Herouville-Saint-Clair, France). The dentine slices appeared as a brightshiny surface littered with dark resorption pits. The softwarecalculated the resorption areas for each slice. When determining theeffects of the compounds in co-cultures, it was necessary to use boththe values obtained for slices removed at the time when the drugs wereadded (e.g., Day 7), as well as the controls from the end of the study(e.g., Day 10).

Rabbit Osteoclast Culture System

The second model system was the rabbit osteoclast system, where mature,functional osteoclasts were isolated from the long bones of rabbits andcultured on dentine slices.

Osteoclast Isolation. Osteoclasts were isolated from the long bones of2-10 day-old rabbits, as described previously (see, e.g., referenceCoxon et al., 2000). All 4 limbs were removed from the rabbits andplaced in ice-cold PBS. Soft tissue and cartilage were removed and thebones transferred into fresh PBS. The bones were minced in aMEM (withoutFCS), using a scalpel. All the medium and fragments were transferred toa 50 ml tube, vortexed for 3×10 seconds and left to stand for 1 minute.The supernatant was removed and made up to 50 ml/rabbit with medium andFCS so as to give a final concentration of 10% FCS.

Osteoclast Plating. The cells were plated onto dentine slices in a 96well plate, at 100 μL/well (medium: αMEM supplemented with 10% FCS andpenicillin and streptomycin) and left for 4 hours to allow adherence tothe dentine. After this period the medium was removed, and with it thenon-adherent cells. Fresh medium was then added. The remainingpopulation was highly enriched in osteoclasts.

Culturing. At this point, test compounds to be studied were added andthe cells cultured at 37° C. in 5% CO₂ for 48 hours. At the end of theculture, the osteoclasts were identified by staining fortartrate-resistant acid phosphatase (TRAcP) staining. A good number ofosteoclasts in the controls was 100-200.

The results were expressed as a % of the average number of osteoclastsseen in the controls. The resorption pit area was quantified byreflected light microscopy as described previously (see, e.g., van't Hof& Ralston, 1997) and again the results expressed as a % of the controlvalues.

Biological Data

1,4-butanediol was tested for its ability to inhibit osteoclastformation and activity in the murine co-culture system; it had nodetected effect on the osteoclasts.

A number of monoesters of alkane diols were prepared, and IC₅₀ valuesfor macrophage J774 viability assays for many of these are summarised inTable 1. The most potent compounds, ABD-0056 (4BP) and ABD-0085 (10F),are almost an order of magnitude more active than any of the othercompounds studied.

TABLE 1 IC₅₀ (μM) # Compound MTT Alamar Blue 1 ABD-0006 4A >100 — 2ABD-0007 4Bu >100 — 3 ABD-0008 4MB >100 — 4 ABD-0009 4B >100 — 5ABD-0014 4P >100 — 6 ABD-0017 6P >100 — 7 ABD-0019 4C >100 — 8 ABD-00284T 20 30 9 ABD-0030 5T 40 — 10 ABD-0031 6T 40 — 11 ABD-0032 4N >100 — 12ABD-0033 4H >100 — 13 ABD-0034 4PT >100 — 14 ABD-0035 BuI 72 — 15ABD-0036 4I 52 — 16 ABD-0037 3I 72 — 17 ABD-0038 5I 68 — 18 ABD-0039 6I72 — 19 ABD-0040 BuBPA >100 — 20 ABD-0041 3BPA 75 — 21 ABD-0042 4BPA 4545 22 ABD-0043 5BPA >100 — 23 ABD-0044 6BPA 28 — 24 ABD-0049 4BP-Ac — 5025 ABD-0050 4BP-Ome — >100 26 ABD-0053 BuBP >100 — 27 ABD-0054 6BP 35 —28 ABD-0055 5BP 65 — 29 ABD-0056 4BP 3.5 18 30 ABD-0057 3BP >100 — 31ABD-0059 4BPX >100 — 32 ABD-0069 4IB — 90 33 ABD-0070 Me-4BP — >100 34ABD-0072 HO-4BP — 45 35 ABD-0077 5F — 25 36 ABD-0085 10F — 4 37 ABD-00864BP-Br — >100 38 ABD-0087 4BP-NO₂ — 30 39 ABD-0088 4xNO₂-BP — 35 40ABD-0089 Xyl — 9 41 ABD-0090 PBP — 45 42 ABD-0094 Et-4BP — >100 43ABD-0095 BP-4F — 40 44 ABD-0096 DBP-4F — — 45 ABD-0097 4OMe-BP — >100 46ABD-0098 2NO₂-BP — 17 47 ABD-0099 2F-BP — 14 48 ABD-0100 4F-BP — 14 49ABD-0102 4Br-BP — 14 50 ABD-0106 2,3,6-FB — >100 51 ABD-0107 3,4-FB— >100 52 ABD-0108 2,3,4-FB — >100 53 ABD-0109 2,4,5-FB — >100 54ABD-0110 2,4-FB — >100 55 ABD-0111 D2,4-FB — >100

Some compounds were also evaluated using the murine co-culture systemand rabbit osteoclasts.

At a concentration of 100 μM, all of the biphenylcarboxy (BP), trityl(T), and ibuprofenyl (I) compounds tested were found to be potentinhibitors of osteoclast formation and activity in the murine co-culturesystem, as illustrated in FIG. 2.

FIG. 3 shows that at 100 μM, survival of macrophage J774 cells iscomparable to that of genuine osteoclasts. All compounds which wereeffective in macrophages were also effective in the murine co-culture,and no compounds were found to be solely effective in one system. Onlyone compound, ABD-0057 (3BP), showed a significant difference inresponse between the two systems.

Some compounds were also evaluated over a range of concentrations, usingthe MTT and Alamar Blue macrophage J774 viability assays.

The results for the biphenylcarboxy compounds, ABD-0053 (BuBP), ABD-0057(3BP), ABD-0056 (4BP), ABD-0055 (5BP), and ABD-0054 (6BP), using the MTTmacrophage J774 viability assay, are illustrated in FIG. 4.

Compound ABD-0056 (4BP) is the most active of the BP compounds, with anIC₅₀ of 3.5 μM. This is substantially lower than any of the other BPderivatives, or indeed any other derivatives yet encountered (other thanABD-0085 (10F)). The next most active BP compound is ABD-0054 (6BP). Thebutanol compound ABD-0053 (BuBP) (which lacks a terminal hydroxyl group)showed very little activity, suggesting that the biphenylcarboxy (BP)group itself is not toxic. The biphenylcarboxylic acid was not active.

The results for the ibuprofenyl (I) compounds, ABD-0035 (Bul), ABD-0037(31), ABD-0036 (41), ABD-0038 (51), and ABD-0039 (61), using the MTTmacrophage J774 viability assay, are illustrated in FIG. 5.

Again the butanediol derivative, compound ABD-0036 (41), was the mostactive, but by a less significant margin. But note that ibuprofen itselfmay be having a significant effect on the proliferation and survival ofthe macrophages.

The results for various trityl compounds, ABD-0028 (4T), ABD-0030 (5T),and ABD-0031 (6T), using the MTT macrophage J774 viability assay, areillustrated n FIG. 6.

All of the trityl compounds were found to inhibit macrophage function toa high degree, suggesting the high toxicity of the triphenylacetylgroup. The butanediol derivative, compound ABD-0028 (4T), is more potentthan compound ABD-0030 (5T) and compound ABD-0031 (6T).

The results for various biphenylacetyl (BPA) compounds, ABD-0040(BuBPA), ABD-0041 (3BPA), ABD-0042 (4BPA), ABD-0043 (5BPA), and ABD-0044(6BPA), using the MTT macrophage J774 viability assay, are illustratedin FIG. 7.

Again, the butanediol derivative, compound ABD-0042 (4BPA), is veryactive, as is the hexanediol derivative, compound ABD-0044 (6BPA). SinceBPA (4-biphenyl acetic acid; also known as Felbinac), like ibuprofen, isa COX inhibitor, some of the effects may be due to suppression ofprostaglandin synthesis.

The results for various butanediol derivatives, ABD-0042 (4BPA),ABD-0028 (4T), ABD-0056 (4BP), ABD-0036 (41), using the MTT macrophageJ774 viability assay, are also illustrated in FIG. 8.

The results for various butanediol derivatives, ABD-0042 (4BPA),ABD-0028 (4T), ABD-0056 (4BP), ABD-0036 (41), using the Alamar Bluemacrophage J774 viability assay, are also illustrated in FIG. 9.

The results for various substituted biphenyl (BP) compounds, ABD-0098(“2NO2”), ABD-0100 (“4F”), ABD-0099 (“2F”), ABD-0089 (“Xyl”), andABD-0102 (“4Br”), using the Alamar Blue macrophage J774 viability assay,are illustrated in FIG. 10.

The results for various substituted biphenyl (BP) compounds, ABD-0072(“OH”), ABD-0089 (“Dimethyl”), ABD-0070 (“Methyl”), ABD-0094 (“Ethyl”),and ABD-0097 (“Methoxy”), using the Alamar Blue macrophage J774viability assay, are illustrated in FIG. 11.

The results for two fluoro-substituted phenyl compounds, ABD-0085(“10F”) and ABD-0077 (“5F”), using the Alamar Blue macrophage J774viability assay, are illustrated in FIG. 12.

Various butanediol derivatives (ABD-0053 (BuBP), ABD-0057 (3BP),ABD-0056 (4BP), ABD-0055 (5BP), ABD-0054 (6BP)) were further evaluatedusing the murine co-culture system and the rabbit osteoclast culturesystem, and osteoclast number and resorption pit area recorded as afunction of compound concentration.

The results for osteoclast number for the rabbit osteoclast culturesystem are illustrated in FIG. 13.

The results for resorption pit area for the rabbit osteoclast culturesystem are illustrated in FIG. 14.

It was possible to investigate the effects of compounds upon theosteoclast precursors by adding the test compounds at the Day 2 stage ofthe co-culture procedure. The test compounds were added and left incontact with the cell culture for 2 days. A 100% medium change was thenperformed, as it is desirable to remove all of the test compound. Thismust be done very carefully without touching or disturbing the celllayer. Were any compound to remain then it would complicate the resultsby potentially killing osteoclasts as well as their precursors and notgive an accurate indication of the toxicity specifically towardsprecursors.

The results for osteoclast number for the murine co-culture system,where test compound was added at Day 2, removed at Day 4 and incubationcontinued until Day 10 (osteoclast precursors), are illustrated in FIG.15.

The results for resorption pit area for the murine co-culture system,where test compound was added at Day 2, removed at Day 4 and incubationcontinued until Day 10 (osteoclast precursors), are illustrated in FIG.16.

FIG. 15 shows that ABD-0056 (4BP) is even more effective againstosteoclast precursor formation than against mature osteoclasts. As apotential treatment for disorders involving excess bone removal, thissuggests ABD-0056 (4BP) is a very potent drug capable of stronglyinhibiting the formation of osteoclasts, and then demonstrating a highdegree of toxicity towards those that do develop. The same pattern isdemonstrated by studies on the levels of osteoclast-induced resorption,as shown in FIG. 16. At all concentrations tested, there is virtually noresorption seen.

The results for osteoclast number for the murine co-culture system,where test compound was added at Day 7, and incubation continued untilDay 10 (mature osteoclasts), are illustrated in FIG. 17.

The results for resorption pit area for the murine co-culture system,where test compound was added at Day 7, and incubation continued untilDay 10 (mature osteoclasts), are illustrated in FIG. 18.

Compounds ABD-0056 (4BP) and ABD-0054 (6BP) significantly reduced rabbitosteoclast numbers and resorption activity. Additionally, ABD-0056 (4BP)showed a significant effect at concentrations down to 10 μM. CompoundsABD-0056 (4BP) and ABD-0054 (6BP) also significantly inhibitedosteoclast formation in the murine co-culture system.

The effects on resorption pit area were more pronounced in the murineco-culture than in the rabbit osteoclast system. In the rabbit system,mature, resorbing osteoclasts are present from the start and may resorbbone for some time, until the compound starts affecting the cells. Theco-culture system, however, depends on the actual formation ofosteoclasts from non-resorbing precursors.

The results for osteoclast number and resorption pit area for the murineco-culture system, where test compound (ABD-0056) (4BP) was added at Day7, and incubation continued until Day 10 (mature osteoclasts), areillustrated in FIG. 19.

The results show that the 1,4-butanediol ester of biphenylcarboxylicacid (compound ABD-0056 (4BP)) is a potent inhibitor of osteoclastsurvival, formation and activity, and is 10 times more effective thanany compounds of this class (other than ABD-0085 (10F)).

In a comparable study on J774 survival (see, e.g., reference Luckman etal., 1998), the bisphophonates alendronate and pamidronate had IC50values that were 10 and 8 times higher respectively.

Without wishing to be bound by any particular theory, it is believedthat the biphenylcarboxylic acid conveys the lipophilicity needed forthe compound to pass across the cell membrane.

Additional esters of 1,4-butanediol were prepared and tested,specifically, the naphth-1-ylacetyl (compound ABD-0032 (4N)),homoveratryl (compound ABD-0033 (4H)), 2-biphenylcarboxy (compoundABD-0059 (4BPX)) and 4-phenyltoluyl (compound ABD-0034 (4PT))derivatives. These compounds showed little activity at 100 μMconcentration.

Without wishing to be bound to any particular theory, it is believedthat the activity of the compounds is related to the overall shape ofthe molecule. The I, BP and BPA derivatives can be regarded as broadlylinear, whereas the BPX and N derivatives are bent out of the linearplane. That the H and PT derivatives are inactive suggests that themolecular target will only accommodate a specific size of compound, andthat the homoveratrate compound is too small or insufficientlylipophilic and the phenyltoluate compound is too long or too flexible.

It is believed that one preferred class of compounds will be of agenerally rigid, cylindrical shape, and be based on the biphenylstructure. A sub-class of such compounds are those with a further4′-substituent on the biphenyl group.

Another sub-class of such compounds are those with further 2′- and4′-substituents on the biphenyl group. Another sub-class of suchcompounds are those with further 2-, 2′- and 4′-substituents on thebiphenyl group.

The foregoing has described the principles, preferred embodiments, andmodes of operation of the present invention. However, the inventionshould not be construed as limited to the particular embodimentsdiscussed. Instead, the above-described embodiments should be regardedas illustrative rather than restrictive, and it should be appreciatedthat variations may be made in those embodiments by workers skilled inthe art without departing from the scope of the present invention asdefined by the appended claims.

REFERENCES

A number of patents and publications are cited above in order to morefully describe and disclose the invention and the state of the art towhich the invention pertains. Full citations for these references areprovided below. Each of these references is incorporated herein byreference in its entirety into the present disclosure, to the sameextent as if each individual reference was specifically and individuallyindicated to be incorporated by reference.

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1. A compound selected from compounds of the following formula:

wherein: A is —(CH₂)_(n)— where n is an integer from 4 to 8; R^(A1) is independently a group of the following formula:

wherein q is an integer from 0 to 4, r is an integer from 0 to 5, and each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —C(═O)OH, —C(═O)OMe, —C(═O)OEt, —C(═O)O(tBu), —C(═O)OPh, —OC(C═O)Me, —OC(C═O)Et, —OC(C═O)(tBu), —OC(C═O)Ph, —OC(C═O)OMe, —OC(C═O)OEt, —OC(C═O)O(tBu), —OC(C═O)OPh, —C(═O)NH₂, —C(═O)NHMe, —C(═O)NMe₂, —C(═O)NHPh, —NHC(═O)Me, —NHC(═O)Et, —NHC(═O)Ph, —NH₂, —NHme, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh; and pharmaceutically acceptable salts thereof.
 2. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


3. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:

wherein s is an integer from 0 to
 4. 4. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


5. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:

wherein s is an integer from 0 to
 4. 6. A compound according to claim 1, wherein R^(A1) is independently group of the following formula:


7. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:

wherein t is an integer from 0 to
 3. 8. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


9. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:

wherein s is an integer from 0 to
 4. 10. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


11. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:

wherein t is an integer from 0 to
 3. 12. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


13. A compound according to claim 1, wherein R^(A1) is independently a group of the following formula:


14. A compound according to claim 2, wherein A is a group of the formula —(CH₂)₄—.
 15. A compound according to claim 2, wherein A is a group of the formula —(CH₂)₅—.
 16. A compound according to claim 2, wherein A is a group of the formula —(CH₂)₆—.
 17. A compound according to claim 3, wherein A is a group of the formula —(CH₂)₄—.
 18. A compound according to claim 3, wherein A is a group of the formula —(CH₂)₅—.
 19. A compound according to claim 3, wherein A is a group of the formula —(CH₂)₆—.
 20. A compound according to claim 4, wherein A is a group of the formula —(CH₂)₄—.
 21. A compound according to claim 4, wherein A is a group of the formula —(CH₂)₅—.
 22. A compound according to claim 4, wherein A is a group of the formula —(CH₂)₆—.
 23. A compound according to claim 5, wherein A is a group of the formula —(CH₂)₄—.
 24. A compound according to claim 5, wherein A is a group of the formula —(CH₂)₅—.
 25. A compound according to claim 5, wherein A is a group of the formula —(CH₂)₆—.
 26. A compound according to claim 9, wherein A is a group of the formula —(CH₂)₄—.
 27. A compound according to claim 9, wherein A is a group of the formula —(CH₂)₅—.
 28. A compound according to claim 9, wherein A is a group of the formula —(CH₂)₆—.
 29. A compound according to claim 10, wherein A is a group of the formula —(CH₂)₄—.
 30. A compound according to claim 10, wherein A is a group of the formula —(CH₂)₅—.
 31. A compound according to claim 10, wherein A is a group of the formula —(CH₂)₆—.
 32. A compound according to claim 11, wherein A is a group of the formula —(CH₂)₄.
 33. A compound according to claim 11, wherein A is a group of the formula —(CH₂)₅—.
 34. A compound according to claim 11, wherein A is a group of the formula —(CH₂)₆—.
 35. A compound according to claim 12, wherein A is a group of the formula —(CH₂)₄—.
 36. A compound according to claim 12, wherein A is a group of the formula —(CH₂)₅—.
 37. A compound according to claim 12, wherein A is a group of the formula —(CH₂)₆—.
 38. A compound according to claim 14, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 39. A compound according to claim 15, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe —S(═O)₂OEt and —S(═O)₂OPh.
 40. A compound according to claim 16, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe —S(═O)₂OEt and —S(═O)₂OPh.
 41. A compound according to claim 17, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 42. A compound according to claim 18, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 43. A compound according to claim 19, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 44. A compound according to claim 20, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 45. A compound according to claim 21, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 46. A compound according to claim 22, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 47. A compound according to claim 23, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 48. A compound according to claim 24, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 49. A compound according to claim 25, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 50. A compound according to claim 26, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 51. A compound according to claim 27, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 52. A compound according to claim 28, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 53. A compound according to claim 29, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 54. A compound according to claim 30, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 55. A compound according to claim 31, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 56. A compound according to claim 32, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 57. A compound according to claim 33, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 58. A compound according to claim 34, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 59. A compound according to claim 35, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 60. A compound according to claim 36, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂—NHMe, —NMe₂—NHEt, —NEt₂, —NO₂—CN, —S(═O)₂OMe, —S(═O)₂OEt, and —S(═O)₂OPh.
 61. A compound according to claim 37, wherein each R^(P) is independently selected from: -Me, -Et, -iPr, -nPr, -tBu, -Ph, —F, —Cl, —Br, —I, —OH, —NH₂, —NHMe, —NMe₂, —NHEt, —NEt₂, —NO₂, —CN, —S(═O)₂OMe, —S(═O)₂OEt and —S(═O)₂OPh.
 62. A compound according to claim 14, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 63. A compound according to claim 15, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 64. A compound according to claim 16, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 65. A compound according to claim 17, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 66. A compound according to claim 18, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 67. A compound according to claim 19, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 68. A compound according to claim 20, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 69. A compound according to claim 21, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 70. A compound according to claim 22, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 71. A compound according to claim 23, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 72. A compound according to claim 24, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 73. A compound according to claim 25, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 74. A compound according to claim 26, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 75. A compound according to claim 27, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 76. A compound according to claim 28, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 77. A compound according to claim 29, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 78. A compound according to claim 30, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 79. A compound according to claim 31, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 80. A compound according to claim 32, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 81. A compound according to claim 33, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 82. A compound according to claim 34, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 83. A compound according to claim 35, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 84. A compound according to claim 36, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 85. A compound according to claim 37, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, and —CN.
 86. A compound according to claim 1, wherein the compound is selected from compounds of the following formulae, and pharmaceutically acceptable salts thereof:


87. A compound according to claim 1, wherein the compound is selected from compounds of the following formula, and pharmaceutically acceptable salts thereof:


88. A compound according to claim 1, wherein the compound is selected from compounds of the following formula:


89. A composition comprising a compound according to claim 1 and a pharmaceutically acceptable carrier or diluent.
 90. A composition comprising a compound according to claim 2 and a pharmaceutically acceptable carrier or diluent.
 91. A composition comprising a compound according to claim 62 and a pharmaceutically acceptable carrier or diluent.
 92. A composition comprising a compound according to claim 86 and a pharmaceutically acceptable carrier or diluent.
 93. A composition comprising a compound according to claim 3 and a pharmaceutically acceptable carrier or diluent.
 94. A composition comprising a compound according to claim 87 and a pharmaceutically acceptable carrier or diluent.
 95. A composition comprising a compound according to claim 88 and a pharmaceutically acceptable carrier or diluent.
 96. A compound according to claim 2, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, and —I.
 97. A compound according to claim 3, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, and —I.
 98. A compound according to claim 14, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, and —I.
 99. A compound according to claim 17, wherein each R^(P) is independently selected from: -Me, —F, —Cl, —Br, and —I.
 100. A compound selected from compounds of the following formula:

wherein: A is —(CH₂)₄—, —(CH₂)₅—, or —(CH₂)₆—; R^(A1) is independently a group of the following formula:

wherein: q is 0 or 1; r is 0, 1, or 2; and each R^(P) is independently -Me, —F, —Cl, —Br, —I, —OH, —NH₂, —NMe₂, —NO₂, or —CN; and pharmaceutically acceptable salts thereof. 